Zebrafish dscaml1 Deficiency Impairs Retinal Patterning and Oculomotor Function.

Down syndrome cell adhesion molecules (dscam and dscaml1) are essential regulators of neural circuit assembly, but their roles in vertebrate neural circuit function are still mostly unexplored. We investigated the functional consequences of dscaml1 deficiency in the larval zebrafish (sexually undifferentiated) oculomotor system, where behavior, circuit function, and neuronal activity can be precisely quantified. Genetic perturbation of dscaml1 resulted in deficits in retinal patterning and light adaptation, consistent with its known roles in mammals. Oculomotor analyses revealed specific deficits related to the dscaml1 mutation, including severe fatigue during gaze stabilization, reduced saccade amplitude and velocity in the light, greater disconjugacy, and impaired fixation. Two-photon calcium imaging of abducens neurons in control and dscaml1 mutant animals confirmed deficits in saccade-command signals (indicative of an impairment in the saccadic premotor pathway), whereas abducens activation by the pretectum-vestibular pathway was not affected. Together, we show that loss of dscaml1 resulted in impairments in specific oculomotor circuits, providing a new animal model to investigate the development of oculomotor premotor pathways and their associated human ocular disorders.SIGNIFICANCE STATEMENTDscaml1 is a neural developmental gene with unknown behavioral significance. Using the zebrafish model, this study shows that dscaml1 mutants have a host of oculomotor (eye movement) deficits. Notably, the oculomotor phenotypes in dscaml1 mutants are reminiscent of human ocular motor apraxia, a neurodevelopmental disorder characterized by reduced saccade amplitude and gaze stabilization deficits. Population-level recording of neuronal activity further revealed potential subcircuit-specific requirements for dscaml1 during oculomotor behavior. These findings underscore the importance of dscaml1 in the development of visuomotor function and characterize a new model to investigate potential circuit deficits underlying human oculomotor disorders.

STAT3-mediated activation of miR-21 is involved in down-regulation of TIMP3 and neovascularization in the ischemic retina.

Retinal neovascularization (RNV) is a sight threatening complication of ischemic retinopathies with limited therapeutic options. The transcription factor signal transducer and activator of transcription 3 (STAT3) has been shown to play a crucial role in promoting RNV. However, manipulating of STAT3 activity can cause significant adverse side effects due to its neurotrophic properties. In this study, we identified microRNA-21 (miR-21) as a downstream effector of STAT3 activity in the ischemic retinas and determined its role in promoting RNV through inhibition of its molecular target, the tissue inhibitor of matrix metalloproteinases 3 (TIMP3). Using human retinal endothelial cells (HREC) exposed to hypoxia and a mouse model of oxygen-induced retinopathy (OIR), we found that TIMP3 expression was significantly decreased at both mRNA and protein levels and this paralleled the activation of STAT3 and up-regulation of miR-21. Moreover, TIMP3 expression was restored by knockdown of STAT3 or blocking of miR-21 in HREC, thus, confirming TIMP3 as a downstream target of STAT3/miR-21 pathway. Finally, in a mouse model of OIR, blockade of miR-21 by a specific antisense (a.miR-21), halted RNV and this effect was associated with rescuing of TIMP3 expression. Our data show that miR-21 mediates STAT3 pro-angiogenic effects in the ischemic retina, thus suggesting its blockade as a potential therapy to prevent/halt RNV.

Utility of molecular testing for related retinal dystrophies.

BACKGROUND: The purpose of this study was to describe our experience with the clinical effects of molecular genetic testing for retinitis pigmentosa (RP) and related retinal dystrophies. METHODS: Chart review of 303 consecutive patients with retinal dystrophies was done when blood was sent for molecular genetic testing between 1993 and 2001. Phenotype information was retrieved for patients with identified mutations. The yield of positive and clinically useful results was assessed. RESULTS: Participants comprised 35 patients with Leber congenital amaurosis, 18 with Usher syndrome, and 250 with isolated RP or other retinal dystrophies. Of these 303 participants, 203 (67%) received positive or negative results of molecular testing for an average of 2.7 genes. Positive results were available in 19 patients after an average time interval of 38+/-22 months (median 33 months, range 1-89 months). No results were received for 84 (28%) patients. In 16 (5%) cases, patients received partial results. Only 19 (6%) patients were found to have sequence changes in RHO, RDS, CRB1, or USH2A, 2 of which were thought to be disease-causing. Only 2 sequence changes were previously documented mutations, but several other novel changes were suspected to be disease-causing mutations also. INTERPRETATION: Molecular testing was helpful only in the minority of cases, largely because of a lack of availability, as well as the complexity of the molecular genetics of RP. Improvements in funding, infrastructure, and molecular knowledge will be necessary to improve the transformation of molecular genetic testing into a clinically relevant bedside tool.

Systemic effects of ophthalmic cyclopentolate on body weight in neonatal mice.

BACKGROUND: Cyclopentolate is standardly used in ophthalmologic examinations of neonates to facilitate screening for retinopathy of prematurity. Reports of systemic effects have raised concerns of an increased risk of feeding intolerance after the examinations. OBJECTIVES: The goal of this study was to evaluate systemic concentrations of cyclopentolate after ophthalmic administration, as well as assess changes in weight as an indirect measure of alteration in feeding. METHODS: Neonatal mice were randomized into three groups to simulate a neonatal model for ophthalmic medication administration. The cyclopentolate group received a one-time administration of tetracaine, cyclopentolate, and phenylephrine ophthalmologic solutions in accordance with the protocol used at the children's hospital. The placebo group received the same ophthalmic drop administration, except for normal saline in place of cyclopentolate, and the control group received no ophthalmic drops and minimal handling. Daily weights and serum samples to measure systemic concentrations of cyclopentolate post-ophthalmic administration were assessed at baseline and for 7 days following drop administration. RESULTS: Analysis of serum levels demonstrated detectability of systemic cyclopentolate after ophthalmic administration as early as 30 min (86 ng/ml), 1 h (60 ng/ml), and 24 h (6.2 ng/ml). There were also differences in weight gained on following ophthalmic administration observed between the cyclopentolate group and placebo group, with the cyclopentolate group weighing significantly less on days 3 and 7 (p = 0.02). CONCLUSIONS: RESULTS indicate cyclopentolate is absorbed systemically and instillation of cyclopentolate decreases weight gain in neonatal mice compared to placebo. These preclinical findings provide rationale for further studies in neonatal patients.

Eye position compensation improves estimates of response magnitude and receptive field geometry in alert monkeys.

Studies of visual function in behaving subjects require that stimuli be positioned reliably on the retina in the presence of eye movements. Fixational eye movements scatter stimuli about the retina, inflating estimates of receptive field dimensions, reducing estimates of peak responses, and blurring maps of receptive field subregions. Scleral search coils are frequently used to measure eye position, but their utility for correcting the effects of fixational eye movements on receptive field maps has been questioned. Using eye coils sutured to the sclera and preamplifiers configured to minimize cable artifacts, we reexamined this issue in two rhesus monkeys. During repeated fixation trials, the eye position signal was used to adjust the stimulus position, compensating for eye movements and correcting the stimulus position to place it at the desired location on the retina. Estimates of response magnitudes and receptive field characteristics in V1 and in LGN were obtained in both compensated and uncompensated conditions. Receptive fields were narrower, with steeper borders, and response amplitudes were higher when eye movement compensation was used. In sum, compensating for eye movements facilitated more precise definition of the receptive field. We also monitored horizontal vergence over long sequences of fixation trials and found the variability to be low, as expected for this precise behavior. Our results imply that eye coil signals can be highly accurate and useful for optimizing visual physiology when rigorous precautions are observed.