Semistructured Observation of Population-based Eye Screening in The Netherlands.

BACKGROUND: In the Netherlands, youth-healthcare (YHC) physicians screen children 7 times for vision disorders between the ages of 1 and 60 months. Examination consists of inspection of the external structures of the eye, fundus red reflex, Hirschberg test, pupillary reflexes, cover-uncover test, alternating-cover test, eye motility, monocular pursuit, and, from 36 months onwards, visual acuity (VA). We observed how well these tests are done. METHODS: Screening test performance was assessed with semistructured observations. Two orthoptic students developed a semistructured observation form. In addition to extensive instructions from an orthoptist and YHC-physicians instructor, they attended 2 one-day courses for YHC physicians. Tests were assessed using criteria based on the Dutch Child Vision Screening Guideline version 2010 and the Dutch Manual for Orthoptic Examination. Type of chart, testing distance, and starting eye were recorded for VA measurements. The observations in the first week were done simultaneously by the two observers and checked for concordance. RESULTS: Concordance between the two observers was good. Twenty-five YHC physicians were observed during 100 days in total. Two physicians were excluded because they examined few children. The remaining 23 physicians examined 329 children, of whom 82 were aged 1-4 months, 157 aged 6-24 months, and 90 aged 36-45 months of age. Fundus red reflex was performed in 89% of children, Hirschberg test in 88%, pupillary reflexes in 14%, cover-uncover test in 65%, alternating-cover test in 62%, eye motility in 68%, monocular pursuit in 23%, and VA at 36-45 months in 94%. Forty-eight percent of cover-uncover tests, 36% of alternating-cover tests, and 7% of eye motility tests were performed correctly. VA was measured at 3 meters in 2%, others at 5 meters in accordance with the guideline. A picture chart was used instead of the Landolt-C at the age of 45 months in 23%. VA measurements were performed correctly in 89%, fundus red reflex in 89%, and Hirschberg test in 87%. CONCLUSION: Hirschberg test, fundus red reflex, and VA were adequately tested in most cases. Cover-uncover test, alternating-cover test, and eye motility were often performed inadequately. Pupillary reflexes were skipped often as room lights could not be dimmed.

Effect of omission of population-based eye screening at age 6-9 months in the Netherlands.

PURPOSE: To investigate omission of population-based eye screening at age 6-9 months in the Netherlands. METHODS: Prospective population-based consecutive birth cohort study was used. In two consecutive birth cohorts, children were eye screened at 1-2 and 3-4 months, but at general-health screening at 6-9 months, the second cohort was not eye screened, unless anything conspicuous was noted or in case of positive family history. Data were collected from screening records and anonymous questionnaires. Semi-structured daylong observations were made of physicians examining children aged 0-4 years, including children from the cohorts, by two orthoptic students. RESULTS: 58 of 6059 children (0.96%), in the screened, and 48 of 5482 children (0.88%) in the unscreened group were referred to orthoptist or ophthalmologist, mostly for observed strabismus. Amblyopia, all combined with strabismus, was diagnosed in ten screened (0.17%) versus six unscreened children (0.11%). Most physicians found preverbal examinations and decisions to refer difficult. The observations by orthoptic students revealed that cover test, pupillary reflexes, pursuit movements and eye motility were frequently performed inadequately, contrary to the Hirschberg test, at this age. CONCLUSION: The screened and unscreened group differed little regarding the number of children referred and found to have amblyopia. Referral was mostly based on observed strabismus.

Extracellular ATP hydrolysis inhibits synaptic transmission by increasing ph buffering in the synaptic cleft.

Neuronal computations strongly depend on inhibitory interactions. One such example occurs at the first retinal synapse, where horizontal cells inhibit photoreceptors. This interaction generates the center/surround organization of bipolar cell receptive fields and is crucial for contrast enhancement. Despite its essential role in vision, the underlying synaptic mechanism has puzzled the neuroscience community for decades. Two competing hypotheses are currently considered: an ephaptic and a proton-mediated mechanism. Here we show that horizontal cells feed back to photoreceptors via an unexpected synthesis of the two. The first one is a very fast ephaptic mechanism that has no synaptic delay, making it one of the fastest inhibitory synapses known. The second one is a relatively slow (τ≈200 ms), highly intriguing mechanism. It depends on ATP release via Pannexin 1 channels located on horizontal cell dendrites invaginating the cone synaptic terminal. The ecto-ATPase NTPDase1 hydrolyses extracellular ATP to AMP, phosphate groups, and protons. The phosphate groups and protons form a pH buffer with a pKa of 7.2, which keeps the pH in the synaptic cleft relatively acidic. This inhibits the cone Ca²âº channels and consequently reduces the glutamate release by the cones. When horizontal cells hyperpolarize, the pannexin 1 channels decrease their conductance, the ATP release decreases, and the formation of the pH buffer reduces. The resulting alkalization in the synaptic cleft consequently increases cone glutamate release. Surprisingly, the hydrolysis of ATP instead of ATP itself mediates the synaptic modulation. Our results not only solve longstanding issues regarding horizontal cell to photoreceptor feedback, they also demonstrate a new form of synaptic modulation. Because pannexin 1 channels and ecto-ATPases are strongly expressed in the nervous system and pannexin 1 function is implicated in synaptic plasticity, we anticipate that this novel form of synaptic modulation may be a widespread phenomenon.

Chloride currents in cones modify feedback from horizontal cells to cones in goldfish retina.

In neuronal systems, excitation and inhibition must be well balanced to ensure reliable information transfer. The cone/horizontal cell (HC) interaction in the retina is an example of this. Because natural scenes encompass an enormous intensity range both in temporal and spatial domains, the balance between excitation and inhibition in the outer retina needs to be adaptable. How this is achieved is unknown. Using electrophysiological techniques in the isolated retina of the goldfish, it was found that opening Ca(2+)-dependent Cl(-) channels in recorded cones reduced the size of feedback responses measured in both cones and HCs. Furthermore, we show that cones express Cl(-) channels that are gated by GABA released from HCs. Similar to activation of I(Cl(Ca)), opening of these GABA-gated Cl(-) channels reduced the size of light-induced feedback responses both in cones and HCs. Conversely, application of picrotoxin, a blocker of GABA(A) and GABA(C) receptors, had the opposite effect. In addition, reducing GABA release from HCs by blocking GABA transporters also led to an increase in the size of feedback. Because the independent manipulation of Ca(2+)-dependent Cl(-) currents in individual cones yielded results comparable to bath-applied GABA, it was concluded that activation of either Cl(-) current by itself is sufficient to reduce the size of HC feedback. However, additional effects of GABA on outer retinal processing cannot be excluded. These results can be accounted for by an ephaptic feedback model in which a cone Cl(-) current shunts the current flow in the synaptic cleft. The Ca(2+)-dependent Cl(-) current might be essential to set the initial balance between the feedforward and the feedback signals active in the cone HC synapse. It prevents that strong feedback from HCs to cones flood the cone with Ca(2)(+). Modulation of the feedback strength by GABA might play a role during light/dark adaptation, adjusting the amount of negative feedback to the signal to noise ratio of the cone output.

GPR179 is required for depolarizing bipolar cell function and is mutated in autosomal-recessive complete congenital stationary night blindness.

Complete congenital stationary night blindness (cCSNB) is a clinically and genetically heterogeneous group of retinal disorders characterized by nonprogressive impairment of night vision, absence of the electroretinogram (ERG) b-wave, and variable degrees of involvement of other visual functions. We report here that mutations in GPR179, encoding an orphan G protein receptor, underlie a form of autosomal-recessive cCSNB. The Gpr179(nob5/nob5) mouse model was initially discovered by the absence of the ERG b-wave, a component that reflects depolarizing bipolar cell (DBC) function. We performed genetic mapping, followed by next-generation sequencing of the critical region and detected a large transposon-like DNA insertion in Gpr179. The involvement of GPR179 in DBC function was confirmed in zebrafish and humans. Functional knockdown of gpr179 in zebrafish led to a marked reduction in the amplitude of the ERG b-wave. Candidate gene analysis of GPR179 in DNA extracted from patients with cCSNB identified GPR179-inactivating mutations in two patients. We developed an antibody against mouse GPR179, which robustly labeled DBC dendritic terminals in wild-type mice. This labeling colocalized with the expression of GRM6 and was absent in Gpr179(nob5/nob5) mutant mice. Our results demonstrate that GPR179 plays a critical role in DBC signal transduction and expands our understanding of the mechanisms that mediate normal rod vision.

Synaptic transmission from horizontal cells to cones is impaired by loss of connexin hemichannels.

In the vertebrate retina, horizontal cells generate the inhibitory surround of bipolar cells, an essential step in contrast enhancement. For the last decades, the mechanism involved in this inhibitory synaptic pathway has been a major controversy in retinal research. One hypothesis suggests that connexin hemichannels mediate this negative feedback signal; another suggests that feedback is mediated by protons. Mutant zebrafish were generated that lack connexin 55.5 hemichannels in horizontal cells. Whole cell voltage clamp recordings were made from isolated horizontal cells and cones in flat mount retinas. Light-induced feedback from horizontal cells to cones was reduced in mutants. A reduction of feedback was also found when horizontal cells were pharmacologically hyperpolarized but was absent when they were pharmacologically depolarized. Hemichannel currents in isolated horizontal cells showed a similar behavior. The hyperpolarization-induced hemichannel current was strongly reduced in the mutants while the depolarization-induced hemichannel current was not. Intracellular recordings were made from horizontal cells. Consistent with impaired feedback in the mutant, spectral opponent responses in horizontal cells were diminished in these animals. A behavioral assay revealed a lower contrast-sensitivity, illustrating the role of the horizontal cell to cone feedback pathway in contrast enhancement. Model simulations showed that the observed modifications of feedback can be accounted for by an ephaptic mechanism. A model for feedback, in which the number of connexin hemichannels is reduced to about 40%, fully predicts the specific asymmetric modification of feedback. To our knowledge, this is the first successful genetic interference in the feedback pathway from horizontal cells to cones. It provides direct evidence for an unconventional role of connexin hemichannels in the inhibitory synapse between horizontal cells and cones. This is an important step in resolving a long-standing debate about the unusual form of (ephaptic) synaptic transmission between horizontal cells and cones in the vertebrate retina.

Hemichannel-mediated and pH-based feedback from horizontal cells to cones in the vertebrate retina.

BACKGROUND: Recent studies designed to identify the mechanism by which retinal horizontal cells communicate with cones have implicated two processes. According to one account, horizontal cell hyperpolarization induces an increase in pH within the synaptic cleft that activates the calcium current (Ca(2+)-current) in cones, enhancing transmitter release. An alternative account suggests that horizontal cell hyperpolarization increases the Ca(2+)-current to promote transmitter release through a hemichannel-mediated ephaptic mechanism. METHODOLOGY/PRINCIPAL FINDINGS: To distinguish between these mechanisms, we interfered with the pH regulating systems in the retina and studied the effects on the feedback responses of cones and horizontal cells. We found that the pH buffers HEPES and Tris partially inhibit feedback responses in cones and horizontal cells and lead to intracellular acidification of neurons. Application of 25 mM acetate, which does not change the extracellular pH buffer capacity, does lead to both intracellular acidification and inhibition of feedback. Because intracellular acidification is known to inhibit hemichannels, the key experiment used to test the pH hypothesis, i.e. increasing the extracellular pH buffer capacity, does not discriminate between a pH-based feedback system and a hemichannel-mediated feedback system. To test the pH hypothesis in a manner independent of artificial pH-buffer systems, we studied the effect of interfering with the endogenous pH buffer, the bicarbonate/carbonic anhydrase system. Inhibition of carbonic anhydrase allowed for large changes in pH in the synaptic cleft of bipolar cell terminals and cone terminals, but the predicted enhancement of the cone feedback responses, according to the pH-hypothesis, was not observed. These experiments thus failed to support a proton mediated feedback mechanism. The alternative hypothesis, the hemichannel-mediated ephaptic feedback mechanism, was therefore studied experimentally, and its feasibility was buttressed by means of a quantitative computer model of the cone/horizontal cell synapse. CONCLUSION: We conclude that the data presented in this paper offers further support for physiologically relevant ephaptic interactions in the retina.

Lateral gain control in the outer retina leads to potentiation of center responses of retinal neurons.

The retina can function under a variety of adaptation conditions and stimulus paradigms. To adapt to these various conditions, modifications in the phototransduction cascade and at the synaptic and network levels occur. In this paper, we focus on the properties and function of a gain control mechanism in the cone synapse. We show that horizontal cells, in addition to inhibiting cones via a "lateral inhibitory pathway," also modulate the synaptic gain of the photoreceptor via a "lateral gain control mechanism." The combination of lateral inhibition and lateral gain control generates a highly efficient transformation. Horizontal cells estimate the mean activity of cones. This mean activity is subtracted from the actual activity of the center cone and amplified by the lateral gain modulation system, ensuring that the deviation of the activity of a cone from the mean activity of the surrounding cones is transmitted to the inner retina with high fidelity. Sustained surround illumination leads to an enhancement of the responses of transient ON/OFF ganglion cells to a flickering center spot. Blocking feedback from horizontal cells not only blocks the lateral gain control mechanism in the outer retina, but it also blocks the surround enhancement in transient ON/OFF ganglion cells. This suggests that the effects of the outer retinal lateral gain control mechanism are visible in the responses of ganglion cells. Functionally speaking, this result illustrates that horizontal cells are not purely inhibitory neurons but have a role in response enhancement as well.