Membrane flow during pinocytosis. A stereologic analysis.

HRP has been used as a cytochemical marker for a sterelogic analysis of pinocytic vesicles and secondary lysosomes in cultivated macrophages and L cells. Evidence is presented that the diaminobenzidine technique (a) detects all vaculoes containing encyme and (b) distinguishes between incoming pinocytic vesicles and those which have fused with pre-existing lysosomes to form secondary lososomes. The HRP reactive pinocytic vesicle spaces fills completely within 5 min after exposure to enzyme, while the secondary lysosome compartment is saturated in 45--60 min. The size distribution of sectioned (profile) vaculoe diameters was measured at equilibrium and converted to actual (spherical) dimensions using a technique modified from Dr. S. D. Wicksell. The most important findings in this study have to do with the rate at which pinocytosed fluid and surface membrane move into the cell and on their subsequent fate. Each minute macrophages form at least 125 pinocytic vesicles having a fractional vol of 0.43% of the cell's volume and a fractional area of 3.1% of the cell's surface area. The fractional volume and surface area flux rates for L cells were 0.05% and 0.8% per minute respectively. Macrophages and L cells thus interiorize the equivalent of their cell surface area every 33 and 125 min. During a 3-period, the size of the secondary lysosome compartment remains constant and represents 2.5% of the cell volume and 18% of the surface area. Each hour, therefore, the volume and surface area of incoming vesicles is 10 times greater than the dimensions of the secondary lysosomes in both macrophages and L cells. This implies a rapid reduction in vesicle size during the formation of the secondary lysosome and the egress of pinocytosed fluid from the vacuole and the cell. In addition, we postulate that membrane components of the vacuole are subsequently recycled back to the cell surface.

The response of the Limulus retina to moving stimuli: a prediction by Fourier synthesis.

The Limulus retina responds as a linear system to light stimuli which vary moderately about a mean level. The dynamics of such a system may conveniently be summarized by means of a spatiotemporal transfer function, which describes the response of the system to moving sinusoidal gratings. The response of the system to an arbitrary stimulus may then be calculated by adding together the system's responses to suitably weighted sinusoidal stimuli. We have measured such a spatiotemporal transfer function for the Limulus eye. We have then accurately predicted, in a parameter-free calculation, the eye's response to various stimulus patterns which move across it at several different velocities.

The spatiotemporal transfer function of the Limulus lateral eye.

The dynamics of the Limulus retina may be well described by the spatiotemporal transfer function, which measures the response of the eye to moving sinusoidal gratings. We consider a model for this system, which incorporates an excitatory generator potential, and self- and lateral inhibitory processes. Procedures are described which allow estimation of parameters for the model consistent with the empirical transfer function data. Transfer functions calculated from the model show good agreement with laboratory measurements, and may be used to predict accurately the response of the eye to arbitrary moving stimuli. The model allows convenient interpretation of the transfer function measurements in terms of physiological processes which underly the response of the Limulus retina.

Photographic calibration of the Hirschberg test.

A series of direct photographic measurements of corneal reflex positions was taken to reconcile discrepant interpretations of the Hirschberg test. A value of 21 prism diopters per millimeter was obtained for the conversion factor relating ocular rotations to reflex displacement, valid over a range of 200 prism D. Quantitative interpretation of routine clinical photographs of strabismic patients can thus be easily obtained by including a millimeter scale in each picture. This value for the conversion factor is in agreement with a simple optical model for the corneal reflex. The discrepancy with the traditional value (about 14 prism D/mm) appears to arise from an attempt of most observers to measure reflex displacement from the corneal apex along the surface of the cornea, rather than to project the reflex and pupil onto a true frontal plane, as is done in a photograph.

An indication for "granny glasses".

A young woman with a large astigmatic refractive error obtained no visual improvement with glasses. Repeated manifest refractions revealed persistent variations in the apparent cylinder axis. A suitable choice of spectacle frames facilitated a satisfactory outcome. The pitfalls inherent in the clinical specification of cylinder axis, and the potential visual consequences, are discussed.

Retinal dysfunction as an initial ophthalmic sign in AIDS.

Three cases of patients with AIDS in whom severe retinal dysfunction preceded the development of widespread funduscopic abnormalities are presented. The disparity between the minimal extent of visible retinal lesions and the severe loss of retinal function was confirmed by visual field and electroretinographic studies. This pattern of early visual loss in AIDS patients may represent infection of the retina by the HIV virus.

Evaluation of different recording parameters to establish a standard for flash electroretinography in rodents.

Different electrodes and stimulus protocols commonly used for electroretinography in rodent eyes were compared for convenience of use, degree of damage to corneal epithelium, and for magnitude of amplitude, reproducibility, left versus right eye accuracy, and reliability of recorded parameters of the flash electroretinogram (ERG). Adult C57BL/6 pigmented mice and albino Wistar rats were used to determine scotopic ERGs in response to Ganzfeld or strobe-light stimulation and light-adapted (photopic) ERGs recorded from both eyes at the same time. Test-retest data were used for statistical analyses to compare a monopolar gold-wire contact lens electrode (CLE), a cotton-wick silver-silver chloride electrode (CSCE), a DTL fiber electrode (DTLE), and a circular stainless steel wire electrode (SSE). Corneas were evaluated for abrasion after ERG recordings using fluorescein staining and also for the time taken, ease of insertion, and re-insertions required for the different electrodes. Compared to CSCE, DTLE, and SSE, the ERG potentials recorded by CLE had significantly larger scotopic amplitudes and oscillatory potentials under strobe or Ganzfeld stimulation and for light-adapted ERG b-wave amplitudes in both mice and rats. In analyzing test-retest data of scotopic ERG a-wave and b-wave amplitudes, the intraclass correlation coefficient showed the best agreement for the CLE (range 0.61-0.94) compared to the SSE (0.13-0.77), DTLE (0.02-0.69), and CSCE (0.12-0.51). In mice and rats, logistic regression analyses revealed significant correlations for amplitudes of most scotopic ERG parameters between contralateral eyes obtained with CLE and for some ERG components recorded by SSE. When comparing ERG amplitudes for stimulation by strobe or Ganzfeld, the difference was least with the CLE compared to DTLE, CSCE, or SSE. The time taken to insert the four different electrodes was greatest for the CLE in both mice and rats. The extent of corneal abrasion resulting from electrode use in mice was largest for the SSE followed by the CLE. However, in rats there was almost no corneal damage after ERG recordings with the CLE. Because of the stability of eye contact, the CLE allows ERGs to be determined over a longer recording session. Recording of scotopic and photopic (light-adapted) ERGs in rodents with monopolar gold-wire contact lens electrodes provides greater amplitudes and higher reproducibility when compared to other commonly used corneal electrodes. These electrodes are significantly better overall than others that were evaluated and should be considered for a standard protocol to monitor retinal function in rodent eyes.

[Pilot study of pattern-electroretinographic changes in the DBA/2NNia mouse. Animal model of congenital angle-closure glaucoma]. / Pilotstudie zu musterelektroretinographischen Veränderungen bei der DBA/2NNia-Maus. Tiermodell eines angeborenen Engwinkelglaukoms.

PURPOSE: The aim of this study was to document the time-course of retinal dysfunction by pattern-electroretinography (PERG) in eyes of the DBA/2NNia substrain of mouse that develop an inherited angle-closure glaucoma. METHODS: Twelve DBA/2NNia mice and 12 control C57BL/6 J mice were studied by PERG recordings from 2 to 10 months of age. PERGs were recorded using different spatial and temporal frequencies. RESULTS: PERGs recorded with a temporal frequency of 7.5 Hz and a spatial frequency of 0.4 cycles/degree performed best to discriminate between DBA/2NNia mice and C57BL/6 J mice. When compared with normal C57BL/6 J mice, significant amplitude reductions of the PERG (Student's t-test; p < 0.01) were found in DBA/2NNia mice by 5 months of age and continued to decline as the animals aged. At beginning of follow-up, the mean PERG amplitude in DBA/2NNia mice was 2.3 +/- 0.5 microV. At 5 months of age, the mean PERG amplitude was reduced by 0.9 +/- 0.45 microV (paired t-test; p < 0.0001). CONCLUSION: Previously, a significant loss of retinal ganglion cells was found in the DBA/2NNia mouse substrain at 6-7 months of age. In the present study, we found decreases in PERG amplitudes, occurring from the age of 5 months onward. Similarities with the findings in human glaucoma indicate the relevance of this animal model for further glaucoma research.

Compound heterozygous PNPLA6 mutations cause Boucher-Neuhäuser syndrome with late-onset ataxia.

PNPLA6 mutations, known to be associated with the development of motor neuron phenotypes, have recently been identified in families with Boucher-Neuhäuser syndrome. Boucher-Neuhäuser is a rare autosomal recessive syndrome characterized by the co-occurrence of cerebellar ataxia, hypogonadotropic hypogonadism, and chorioretinal dystrophy. Gait ataxia in Boucher-Neuhäuser usually manifests before early adulthood, although onset in the third or fourth decade has also been reported. However, given the recent identification of PNPLA6 mutations as the cause of this condition, the determining factors of age of symptom onset still need to be established. Here, we have identified a sporadic Boucher-Neuhäuser case with late-onset gait ataxia and relatively milder retinal changes due to compound heterozygous PNPLA6 mutations. Compound heterozygosity was confirmed by cloning and sequencing the patient's genomic DNA from coding exons 26-29. Furthermore, both mutations (one novel and one known) fell in the phospholipase esterase domain, where most pathogenic mutations seem to cluster. Taken together, we herein confirm PNPLA6 mutations as the leading cause of Boucher-Neuhäuser syndrome and suggest inquiring about a history of hypogonadism or visual changes in patients presenting with late-onset gait ataxia. We also advocate for neuroophthalmologic evaluation in suspected cases.

Intra-arterial chemotherapy as a treatment for intraocular retinoblastoma: alternatives to direct ophthalmic artery catheterization.

BACKGROUND AND PURPOSE: Intra-arterial chemotherapy is a very effective treatment option for intraocular retinoblastoma. However, direct catheterization of the OA is not always possible. The purpose of this work was to report our initial results with intra-arterial chemotherapy for intraocular retinoblastoma when delivery of the drug was not via direct catheterization of the OA. MATERIALS AND METHODS: Retrospective review of 110 eyes (89 patients) undergoing a total of 351 intra-arterial treatments at our institution between 2006 and 2010 identified 18 eyes (14 patients) that received at least 1 infusion via a vascular route other than direct OA catheterization. Alternatives included catheterization of the orbital branch of the MMA and temporary balloon occlusion of the ICA. RESULTS: Tumor control was observed in 17 of 18 eyes at a mean follow-up of 18.9 months (median, 17.5 months; range, 8-36 months). The mean number of intra-arterial infusions was 3.7 per eye (median, 3; range, 2-9). Treatment routes included the following: MMA only, 3 eyes; MMA + OA, 4 eyes; MMA + balloon, 2 eyes; balloon only, 1 eye; balloon + OA, 7 eyes; balloon + OA + MMA, 1 eye. Intra-arterial chemotherapies included melphalan, topotecan, and carboplatin. Complications were all transient. ERG readings were the following: stable, 10 eyes; improved, 3 eyes; reduced, 5 eyes. One patient died from a second malignancy (pinealoblastoma). CONCLUSIONS: This initial experience shows that when direct OA catheterization is not possible, using alternative routes of intra-arterial chemotherapy saves eyes and preserves vision with acceptable side effects.

Natural history of Type A Niemann-Pick disease: possible endpoints for therapeutic trials.

OBJECTIVE: To describe the disease course and natural history of Type A Niemann-Pick disease (NPD). METHODS: Ten patients with NPD-A (six male, four female; age range at entry: 3 to 6 months) were serially evaluated including clinical neurologic, ophthalmologic, and physical examinations, and assessment of development. Laboratory analyses, abdominal and brain ultrasounds, and chest radiographs also were obtained and information on intercurrent illnesses and cause of mortality was collected. RESULTS: All affected infants had a normal neonatal course and early development. The first symptom detected in all patients was hepatosplenomegaly. Developmental age did not progress beyond 10 months for adaptive behavior, 12 months for expressive language, 9 months for gross motor skills, and 10 months for fine motor skills. Non-neurologic symptoms included frequent vomiting, failure to thrive, respiratory infections, irritability, and sleep disturbance. Neurologic examination at the time of presentation was normal in most patients. Later neurologic examinations revealed progressive hypotonia with loss of the deep tendon reflexes. All patients had cherry red spots by 12 months. The median time from diagnosis to death was 21 months. The cause of death was respiratory failure in nine patients and complications from bleeding in the tenth. CONCLUSIONS: The clinical course in Type A Niemann-Pick disease is similar among affected patients and is characterized by a relentless neurodegenerative course that leads to death, usually within 3 years.

Corneal topography and the hirschberg test.

A simple trigonometric analysis of the Hirschberg test with the assumption that the corneal surface is spherical predicts a sinusoidal dependence of the corneal reflex displacement on the angle of ocular rotation. A comparison with corneal reflex photographs demonstrates that at angles larger than 50 prism diopters (26 deg) the reflex displacements are larger than predicted by the spherical model. This discrepancy may be accounted for by incorporating a more general description of the corneal topography into the geometric analysis. The linear Hirschberg relation that is seen in typical data is accounted for by a relative flattening of the peripheral cornea by ~ 20% of the apical curvature. This geometric analysis of the functional dependence of the Hirschberg relation on the corneal topography can be expressed as an integral equation. Differentiation yields a second-order differential equation for the corneal topography in terms of the Hirschberg data. If the Hirschberg relation is assumed to be linear, a quadratic dependence is found for the corneal curvature. A similar differential approach can be formulated for the Placido disk. In this sense the corneal topography problem given in terms of Placido disk data is shown to be wellformulated. The relative simplicity of the Hirschberg geometry is seen to stem from the alignment of the light source with the eye of the observer.

Diagnostic yield of the routine dilated fundus examination.

OBJECTIVE: This study aimed to estimate the risk of errors of omission inherent in the elimination of the routine dilated fundus examination in asymptomatic patients presenting for routine eye checkups, or in patients with purely refractive complaints. DESIGN: The study design was a retrospective chart review. PARTICIPANTS: The authors reviewed the records of initial visits of more than 3800 patients drawn from the practices of 3 board-certified ophthalmologists with substantial general ophthalmology components. Patients with best-corrected visual acuity in either eye of poorer than 20/25, myopia greater than -3.00 diopters, or other risk factors for disease of the fundus or optic nerve (such as ocular hypertension, glaucoma, previous ocular surgery, diabetes mellitus, and history of retinal detachment) were excluded. The authors identified 1094 such records that documented the findings of dilated fundus examination, including indirect ophthalmoscopy. MAIN OUTCOME MEASURES: The authors recorded the prevalence of all fundus abnormalities, of those deemed clinically significant, and those beyond the view of routine examination with the direct ophthalmoscope. Findings were stratified by patient age. RESULTS: Of the 1094 eligible records, 53 (4.84%) had recorded fundus abnormalities. Of these, 23 were of no adverse ophthalmologic consequences and were considered clinically insignificant. Of the 30 (2.74%) potentially significant findings, only 3 were located beyond the view of routine examination with the direct ophthalmoscope (0.274%; 95% confidence limits: 0.56%-0.80%). The prevalence of fundus findings increased strongly with increasing patient age, from 2.4% younger than age 20 to 14.8% older than age 60. Clinically significant findings increased from 0.8% younger than age 20 to 8.9% older than age 60. CONCLUSIONS: The authors estimate the rate of detection through routine dilated fundus examination of clinically significant fundus lesions in asymptomatic patients at 2.73% (95% confidence limits, 1.86%-3.80%). One tenth of these are beyond the view of the direct ophthalmoscope. The prevalence of fundus abnormalities increases tenfold with increasing patient age.

Combined amplitude and phase criteria for evaluation of macular electroretinograms.

Focal electroretinograms (ERGs) were obtained from 35 normal eyes and 48 eyes with known macular disease using the MaculoScope stimulator-ophthalmoscope instrument. Response amplitude and phase were combined to permit interpretation of the ERG responses as two-dimensional vectors. This vector interpretation avoids statistical anomalies and distortions inherent in the use of amplitude and phase response parameters. Furthermore, the vector interpretation permits the use of the Hotelling T2 statistic for the assessment of the significance of the differences between ERG responses, providing greater statistical power than simultaneous comparison of amplitude and phase data separately. The distribution of normal responses yielded limiting normal values of 135 nV for the amplitude and -95 degrees for the phase lag of the focal ERG as recorded by the MaculoScope instrument. Significant abnormalities of the focal ERG are illustrated for cases of cilioretinal artery occlusion, Stargardt's disease, idiopathic macular hole, diabetic macular edema, and idiopathic macular dysfunction with normal fundus. This study demonstrates the potential value of the focal ERG in the demonstration of macular disease undetectable by conventional electroretinography.

Treatment of nerve impulse data for comparison with theory.

A procedure is given for the comparison of nerve impulse data with model predictions. This method utilizes information in the nerve impulse train that is ignored by the post-stimulus-onset histogram and thereby gives an improved signal-to-noise ratio. Comparison of observed responses in the Limulus retina with predictions derived from a detailed model gives good agreement.

Effect of boundaries on the response of a neural network.

The effect an abrupt boundary has upon the dynamical response of a neural network is investigated. The retina of the Limulus eye is used as a model system for studying this effect. A theoretical technique is presented for the quantitative prediction of the manner in which this neural network responds in the vicinity of its boundary. Corresponding experimental measurements of the response to moving stimuli by single optic neurons located near retinal boundaries are presented. Theory and experiment show detailed quantitative agreement.