Contribution of the Bony Bankart in Calculating Glenoid Bone Loss.

Background: Determining the magnitude of glenoid bone loss in patients with anterior shoulder instability is an important step in guiding management. Most calculations to estimate the bone loss do not include the bony Bankart fragment. However, if it can be reduced and adequately fixed, the estimation of bone loss may be decreased. Purpose: To derive a simple equation to calculate the surface area of the bony fragment in Bankart fractures. Study Design: Case series; Level of evidence, 4. Methods: A total of 26 patients suspected of having clinically significant bone loss underwent computed tomography imaging preoperatively, and the percentage of glenoid bone loss (%BL) was approximated with imaging software using a freehand region of interest area measurement with and without the inclusion of the bony Bankart fragment. By assuming this bony fragment as a hemi-ellipse with height, H, and thickness, d, we represented the surface are of the bony piece (Abonefragment=πHd4), and subtracted it from the overall %BL. They compared this value with the one found using imaging software. Results: Without the inclusion of the bony Bankart, the overall %BL by the standard true-fit circle measured using imaging software was 23.8% ± 9.7%. When including the bony Bankart, the glenoid %BL measured using imaging software was found to be 12.1% ± 8.5%. The %BL calculated by our equation with the bony Bankart included was 10% ± 11.1%. There was no statistically significant difference between the %BL values measured using the equation and the imaging software (P = .46). Conclusion: Using a simple equation that approximates the bony Bankart fragment as a hemiellipse allowed for estimation of the glenoid bone loss, assuming that the fragment can be reduced and adequately fixed. This method may serve as a helpful tool in preoperative planning when there are considerations for incorporating the bony fragment in the repair.

The Developmental Progression of Eight Opsin Spectral Signals Recorded from the Zebrafish Retinal Cone Layer Is Altered by the Timing and Cell Type Expression of Thyroxin Receptor ß2 (trß2) Gain-Of-Function Transgenes.

Zebrafish retinal cone signals shift in spectral shape through larval, juvenile, and adult development as expression patterns of eight cone-opsin genes change. An algorithm extracting signal amplitudes for the component cone spectral types is developed and tested on two thyroxin receptor ß2 (trß2) gain-of-function lines crx:mYFP-2A-trß2 and gnat2:mYFP-2A-trß2, allowing correlation between opsin signaling and opsin immunoreactivity in lines with different developmental timing and cell-type expression of this red-opsin-promoting transgene. Both adult transgenics became complete, or nearly complete, "red-cone dichromats," with disproportionately large long-wavelength-sensitive (LWS)1 opsin amplitudes as compared with controls, where LWS1 and LWS2 amplitudes were about equal, and significant signals from SWS1, SWS2, and Rh2 opsins were detected. But in transgenic larvae and juveniles of both lines it was LWS2 amplitudes that increased, with LWS1 cone signals rarely encountered. In gnat2:mYFP-2A-trß2 embryos at 5 d postfertilization (dpf), red-opsin immunoreactive cone density doubled, but red-opsin amplitudes (LWS2) increased <10%, and green-opsin, blue-opsin, and UV-opsin signals were unchanged, despite co-expressed red opsins, and the finding that an sws1 UV-opsin reporter gene was shut down by the gnat2:mYFP-2A-trß2 transgene. By contrast both LWS2 red-cone amplitudes and the density of red-cone immunoreactivity more than doubled in 5-dpf crx:mYFP-2A-trß2 embryos, while UV-cone amplitudes were reduced 90%. Embryonic cones with trß2 gain-of-function transgenes were morphologically distinct from control red, blue or UV cones, with wider inner segments and shorter axons than red cones, suggesting cone spectral specification, opsin immunoreactivity and shape are influenced by the abundance and developmental timing of trß2 expression.

Diagnostic Performance and Interreader Agreement of the MRI Clear Cell Likelihood Score for Characterization of cT1a and cT1b Solid Renal Masses: An External Validation Study.

BACKGROUND. The clear cell likelihood score (ccLS) has been proposed for the noninvasive differentiation of clear cell renal cell carcinoma (ccRCC) from other renal neoplasms on multiparametric MRI (mpMRI), though further external validation remains needed. OBJECTIVE. The purpose of our study was to evaluate the diagnostic performance and interreader agreement of the ccLS version 2.0 (v2.0) for characterizing solid renal masses as ccRCC. METHODS. This retrospective study included 102 patients (67 men, 35 women; mean age, 56.9 ± 12.8 [SD] years) who underwent mpMRI between January 2013 and February 2018, showing a total of 108 (≥ 25% enhancing tissue) solid renal masses measuring 7 cm or smaller (83 cT1a [≤ 4 cm] and 25 cT1b [> 4 cm and ≤ 7 cm]), all with a histologic diagnosis. Three abdominal radiologists independently reviewed the MRI examinations using ccLS v2.0. Median reader sensitivity, specificity, and accuracy were computed for predicting ccRCC by ccLS of 4 or greater, and individual reader AUCs were derived. The percentage of masses that were ccRCC was calculated, stratified by ccLS. Interobserver agreement was assessed by the Fleiss kappa statistic. RESULTS. The sample included 45 ccRCCs (34 cT1a, 11 cT1b), 30 papillary renal cell carcinomas (RCCs), 13 chromophobe RCCs, 14 oncocytomas, and six fat-poor angiomyolipomas. Median reader sensitivity, specificity, and accuracy for predicting ccRCC by ccLS of 4 or greater were 85%, 82%, and 83% among cT1a masses and 82%, 100%, and 92% among cT1b masses. The three readers' AUCs for predicting ccRCC by ccLS for cT1a masses were 0.90, 0.84, and 0.89 and for cT1b masses were 0.99, 0.97, and 0.92. Across readers, the percentage of masses that were ccRCC among cT1a masses was 0%, 0%, 20%, 68%, and 93% for ccLS of 1, 2, 3, 4, and 5, respectively; among cT1b masses, the percentage of masses that were ccRCC was 0%, 0%, 32%, 90%, and 100% for ccLS of 1, 2, 3, 4, and 5, respectively. Interobserver agreement among cT1a and cT1b masses for ccLS of 4 or greater was 0.82 and 0.83 and for ccLS of 1-5 overall was 0.65 and 0.62, respectively. CONCLUSION. This study provides external validation of the ccLS, finding overall high measures of diagnostic performance and interreader agreement. CLINICAL IMPACT. The ccLS provides a standardized approach to the noninvasive diagnosis of ccRCC by MRI.

A multiscale continuum model of the vertebrate outer retina: The temporal dynamics of background-induced flicker enhancement.

The retina is a part of the central nervous system that is accessible, well documented, and studied by researchers spanning the clinical, experimental, and theoretical sciences. Here, we mathematically model the subcircuits of the outer plexiform layer of the retina on two spatial scales: that of an individual synapse and that of the scale of the receptive field (hundreds to thousands of synapses). To this end we formulate a continuum spine model (a partial differential equation system) that incorporates the horizontal cell syncytium and its numerous processes (spines) within cone pedicles. With this multiscale modeling approach, detailed biophysical mechanisms at the synaptic level are retained while scaling up to the receptive field level. As an example of its utility, the model is applied to study background-induced flicker enhancement in which the onset of a dim background enhances the center flicker response of horizontal cells. Simulation results, in comparison with flicker enhancement data for square, slit, and disk test regions, suggest that feedback mechanisms that are voltage-axis modulators of cone calcium channels (for example, ephaptic and/or pH feedback) are robust in capturing the temporal dynamics of background-induced flicker enhancement. The value and potential of this continuum spine approach is that it provides a framework for mathematically modeling the input-output properties of the entire receptive field of the outer retina while implementing the latest models for transmission mechanisms at the synaptic level.

Zebrafish Crb1, Localizing Uniquely to the Cell Membranes around Cone Photoreceptor Axonemes, Alleviates Light Damage to Photoreceptors and Modulates Cones' Light Responsiveness.

The crumbs (crb) apical polarity genes are essential for the development and functions of epithelia. Adult zebrafish retinal neuroepithelium expresses three crb genes (crb1, crb2a, and crb2b); however, it is unknown whether and how Crb1 differs from other Crb proteins in expression, localization, and functions. Here, we show that, unlike zebrafish Crb2a and Crb2b as well as mammalian Crb1 and Crb2, zebrafish Crb1 does not localize to the subapical regions of photoreceptors and Müller glial cells; rather, it localizes to a small region of cone outer segments: the cell membranes surrounding the axonemes. Moreover, zebrafish Crb1 is not required for retinal morphogenesis and photoreceptor patterning. Interestingly, Crb1 promotes rod survival under strong white light irradiation in a previously unreported non--cell-autonomous fashion; in addition, Crb1 delays UV and blue cones' chromatin condensation caused by UV light irradiation. Finally, Crb1 plays a role in cones' responsiveness to light through an arrestin-translocation-independent mechanism. The localization of Crb1 and its functions do not differ between male and female fish. We conclude that zebrafish Crb1 has diverged from other vertebrate Crb proteins, representing a neofunctionalization in Crb biology during evolution.SIGNIFICANCE STATEMENT Apicobasal polarity of epithelia is an important property that underlies the morphogenesis and functions of epithelial tissues. Epithelial apicobasal polarity is controlled by many polarity genes, including the crb genes. In vertebrates, multiple crb genes have been identified, but the differences in their expression patterns and functions are not fully understood. Here, we report a novel subcellular localization of zebrafish Crb1 in retinal cone photoreceptors and evidence for its new functions in photoreceptor maintenance and light responsiveness. This study expands our understanding of the biology of the crb genes in epithelia, including retinal neuroepithelium.

Thyroid hormone receptor beta mutations alter photoreceptor development and function in Danio rerio (zebrafish).

We investigate mutations in trß2, a splice variant of thrb, identifying changes in function, structure, and behavior in larval and adult zebrafish retinas. Two N-terminus CRISPR mutants were identified. The first is a 6BP+1 insertion deletion frameshift resulting in a truncated protein. The second is a 3BP in frame deletion with intact binding domains. ERG recordings of isolated cone signals showed that the 6BP+1 mutants did not respond to red wavelengths of light while the 3BP mutants did respond. 6BP+1 mutants lacked optomotor and optokinetic responses to red/black and green/black contrasts. Both larval and adult 6BP+1 mutants exhibit a loss of red-cone contribution to the ERG and an increase in UV-cone contribution. Transgenic reporters show loss of cone trß2 activation in the 6BP+1 mutant but increase in the density of cones with active blue, green, and UV opsin genes. Antibody reactivity for red-cone LWS1 and LWS2 opsin was absent in the 6BP+1 mutant, as was reactivity for arrestin3a. Our results confirm a critical role for trß2 in long-wavelength cone development.

Strain variations in cone wavelength peaks in situ during zebrafish development.

There are four cone morphologies in zebrafish, corresponding to UV (U), blue (B), green (G), and red (R)-sensing types; yet genetically, eight cone opsins are expressed. How eight opsins are physiologically siloed in four cone types is not well understood, and in larvae, cone physiological spectral peaks are unstudied. We use a spectral model to infer cone wavelength peaks, semisaturation irradiances, and saturation amplitudes from electroretinogram (ERG) datasets composed of multi-wavelength, multi-irradiance, aspartate-isolated, cone-PIII signals, as compiled from many 5- to 12-day larvae and 8- to 18-month-old adult eyes isolated from wild-type (WT) or roy orbison (roy) strains. Analysis suggests (in nm) a seven-cone, U-360/B1-427/B2-440/G1-460/G3-476/R1-575/R2-556, spectral physiology in WT larvae but a six-cone, U-349/B1-414/G3-483/G4-495/R1-572/R2-556, structure in WT adults. In roy larvae, there is a five-cone structure: U-373/B2-440/G1-460/R1-575/R2-556; in roy adults, there is a four-cone structure, B1-410/G3-482/R1-571/R2-556. Existence of multiple B, G, and R types is inferred from shifts in peaks with red or blue backgrounds. Cones were either high or low semisaturation types. The more sensitive, low semisaturation types included U, B1, and G1 cones [3.0-3.6 log(quanta·µm-2·s-1)]. The less sensitive, high semisaturation types were B2, G3, G4, R1, and R2 types [4.3-4.7 log(quanta·µm-2·s-1)]. In both WT and roy, U- and B- cone saturation amplitudes were greater in larvae than in adults, while G-cone saturation levels were greater in adults. R-cone saturation amplitudes were the largest (50-60% of maximal dataset amplitudes) and constant throughout development. WT and roy larvae differed in cone signal levels, with lesser UV- and greater G-cone amplitudes occurring in roy, indicating strain variation in physiological development of cone signals. These physiological measures of cone types suggest chromatic processing in zebrafish involves at least four to seven spectral signal processing pools.

Color Processing in Zebrafish Retina.

Zebrafish (Danio rerio) is a model organism for vertebrate developmental processes and, through a variety of mutant and transgenic lines, various diseases and their complications. Some of these diseases relate to proper function of the visual system. In the US, the National Eye Institute indicates >140 million people over the age of 40 have some form of visual impairment. The causes of the impairments range from refractive error to cataract, diabetic retinopathy and glaucoma, plus heritable diseases such as retinitis pigmentosa and color vision deficits. Most impairments directly affect the retina, the nervous tissue at the back of the eye. Zebrafish with long or short-wavelength color blindness, altered retinal anatomy due to hyperglycemia, high intraocular pressure, and reduced pigment epithelium are all used, and directly applicable, to study how these symptoms affect visual function. However, many published reports describe only molecular/anatomical/structural changes or behavioral deficits. Recent work in zebrafish has documented physiological responses of the different cell types to colored (spectral) light stimuli, indicating a complex level of information processing and color vision in this species. The purpose of this review article is to consolidate published morphological and physiological data from different cells to describe how zebrafish retina is capable of complex visual processing. This information is compared to findings in other vertebrates and relevance to disorders affecting color processing is discussed.

Mutation in the intracellular chloride channel CLCC1 associated with autosomal recessive retinitis pigmentosa.

We identified a homozygous missense alteration (c.75C>A, p.D25E) in CLCC1, encoding a presumptive intracellular chloride channel highly expressed in the retina, associated with autosomal recessive retinitis pigmentosa (arRP) in eight consanguineous families of Pakistani descent. The p.D25E alteration decreased CLCC1 channel function accompanied by accumulation of mutant protein in granules within the ER lumen, while siRNA knockdown of CLCC1 mRNA induced apoptosis in cultured ARPE-19 cells. TALEN KO in zebrafish was lethal 11 days post fertilization. The depressed electroretinogram (ERG) cone response and cone spectral sensitivity of 5 dpf KO zebrafish and reduced eye size, retinal thickness, and expression of rod and cone opsins could be rescued by injection of wild type CLCC1 mRNA. Clcc1+/- KO mice showed decreased ERGs and photoreceptor number. Together these results strongly suggest that intracellular chloride transport by CLCC1 is a critical process in maintaining retinal integrity, and CLCC1 is crucial for survival and function of retinal cells.

One month of hyperglycemia alters spectral responses of the zebrafish photopic electroretinogram.

Prolonged hyperglycemia can alter retinal function, ultimately resulting in blindness. Adult zebrafish adults exposed to alternating conditions of 2% glucose/0% glucose display a 3× increase in blood sugar levels. After 4 weeks of treatment, electroretinograms (ERGs) were recorded from isolated, perfused, in vitro eyecups. Control animals were exposed to alternating 2% mannitol/0% mannitol (osmotic control) or to alternating water (0% glucose/0% glucose; handling control). Two types of ERGs were recorded: (1) native ERGs measured using white-light stimuli and medium without synaptic blockers; and (2) spectral ERGs measured with an AMPA/kainate receptor antagonist, isolating photoreceptor-to-ON-bipolar-cell synapses, and a spectral protocol that separated red (R), green (G), blue (B) and UV cone signals. Retinas were evaluated for changes in layer thickness and for the inflammatory markers GFAP and Nf-κB (RelA or p65). In native ERGs, hyperglycemic b- and d-waves were lower in amplitude than the b- and d-waves of mannitol controls. Alteration of waveshape became severe, with b-waves becoming more transient and ERG responses showing more PIII-like (a-wave) characteristics. For spectral ERGs, waveshape appeared similar in all treatment groups. However, a1- and b2-wave implicit times were significantly longer, and amplitudes were significantly reduced, in response to hyperglycemic treatment, owing to the functional reduction in signals from R, G and B cones. Nf-κB increased significantly in hyperglycemic retinas, but the increase in GFAP was not significant and retinal layer thickness was unaffected. Thus, prolonged hyperglycemia triggers an inflammatory response and functional deficits localized to specific cone types, indicating the rapid onset of neural complications in the zebrafish model of diabetic retinopathy.

The role of adenotonsillectomy in the treatment of primary nocturnal enuresis in children: A systematic review.

INTRODUCTION: Primary nocturnal enuresis (PNE) is a challenging condition for physicians, patients and families. Although the etiology remains unclear, sleep-disordered breathing (SDB) and sleep apnea have been suggested to play an important role. Recent research has suggested a potential therapeutic benefit of adenotonsillectomy (T&A) and surgical management of upper airway obstruction in the treatment of PNE. OBJECTIVE: The aim was to conduct a systematic review of relevant literature to determine the effectiveness of T&A in treating children aged 2-19 years with PNE. STUDY DESIGN: This was a systematic review using a comprehensive electronic search strategy that included PubMed, Embase, CINAHL, Cochrane Library, conference proceedings, and the gray literature up to July 2015. We included all studies of children aged 2-19 years with PNE and SDB who underwent T&A. The primary outcome was resolution of PNE following surgery. Observational studies and randomized trials were reviewed. Risk of bias assessment and meta-analyses of included studies were performed. RESULTS: We screened 3254 citations; following title and abstract screening, 42 studies were selected for full-text screening by two independent reviewers. We included 18 studies (890 patients) in our final analysis. All studies were observational and only one included a control group. Meta-analysis of proportions of all (18) studies revealed a pooled complete resolution rate of 51% (43-60%), with significant heterogeneity among studies (I2 = 82.2%). Partial resolution was seen in 20% (14-27%), with similar heterogeneity to the complete resolution group. Sensitivity analysis including only studies with a low risk of bias and with patients ≥5 years (n = 244 patients) yielded a complete resolution rate of 43% (36-49%) with minimal heterogeneity (I2 = 0%; figure). CONCLUSION: In our systematic review, T&A resulted in improvement of nocturnal enuresis in more than 60% of patients, with complete resolution rates in excess of 50%. Findings were persistent on meta-analysis focused only on studies including older patients (≥5 years) and those with short follow-up after surgery (≤3 months), which imply a higher cure rate than would be expected based on natural history alone. The limitations of this review include the lack of controlled trials, the overall quality of the evidence reviewed and the heterogeneity between included studies. The role for systematic investigation and treatment of sleep disorders in patients with PNE should be scrutinized further, since a near 50% complete resolution rate for PNE may be expected with T&A in some settings.

Celsr3 is required for normal development of GABA circuits in the inner retina.

The identity of the specific molecules required for the process of retinal circuitry formation is largely unknown. Here we report a newly identified zebrafish mutant in which the absence of the atypical cadherin, Celsr3, leads to a specific defect in the development of GABAergic signaling in the inner retina. This mutant lacks an optokinetic response (OKR), the ability to visually track rotating illuminated stripes, and develops a super-normal b-wave in the electroretinogram (ERG). We find that celsr3 mRNA is abundant in the amacrine and ganglion cells of the retina, however its loss does not affect synaptic lamination within the inner plexiform layer (IPL) or amacrine cell number. We localize the ERG defect pharmacologically to a late-stage disruption in GABAergic modulation of ON-bipolar cell pathway and find that the DNQX-sensitive fast b1 component of the ERG is specifically affected in this mutant. Consistently, we find an increase in GABA receptors on mutant ON-bipolar terminals, providing a direct link between the observed physiological changes and alterations in GABA signaling components. Finally, using blastula transplantation, we show that the lack of an OKR is due, at least partially, to Celsr3-mediated defects within the brain. These findings support the previously postulated inner retina origin for the b1 component and reveal a new role for Celsr3 in the normal development of ON visual pathway circuitry in the inner retina.

Seasonal regulation of the growth hormone-insulin-like growth factor-I axis in the American black bear (Ursus americanus).

The American black bear maintains lean body mass for months without food during winter denning. We asked whether changes in the growth hormone/insulin-like growth factor-I (GH-IGF-I) axis may contribute to this remarkable adaptation to starvation. Serum IGF-I levels were measured by radioimmunoassay, and IGF-binding proteins (IGFBPs) were analyzed by ligand blotting. Initial studies in bears living in the wild showed that IGF-I levels are highest in summer and lowest in early winter denning. Detailed studies in captive bears showed that IGF-I levels decline in autumn when bears are hyperphagic, continue to decline in early denning, and later rise above predenning levels despite continued starvation in the den. IGFBP-2 increased and IGFBP-3 decreased in early denning, and these changes were also reversed in later denning. Treatment with GH (0.1 mg·kg(-1)·day(-1) × 6 days) during early denning increased serum levels of IGF-I and IGFBP-3 and lowered levels of IGFBP-2, indicating that denning bears remain responsive to GH. GH treatment lowered blood urea nitrogen levels, reflecting effects on protein metabolism. GH also accelerated weight loss and markedly increased serum levels of free fatty acids and ß-hydroxybutyrate, resulting in a ketoacidosis (bicarbonate decreased to 15 meq/l), which was reversed when GH was withdrawn. These results demonstrate seasonal regulation of GH/IGF-I axis activity in black bears. Diminished GH activity may promote fat storage in autumn in preparation for denning and prevent excessive mobilization and premature exhaustion of fat stores in early denning, whereas restoration of GH/IGF activity in later denning may prepare the bear for normal activity outside the den.

Spectral responses in zebrafish horizontal cells include a tetraphasic response and a novel UV-dominated triphasic response.

Zebrafish are tetrachromats with red (R, 570 nm), green (G, 480 nm), blue (B, 415 nm), and UV (U, 362 nm) cones. Although neurons in other cyprinid retinas are rich in color processing neural circuitry, spectral responses of individual neurons in zebrafish retina, a genetic model for vertebrate color vision, are yet to be studied. Using dye-filled sharp microelectrodes, horizontal cell voltage responses to light stimuli of different wavelengths and irradiances were recorded in a superfused eyecup. Spectral properties were assessed both qualitatively and quantitatively. Six spectral classes of horizontal cell were distinguished. Two monophasic response types (L1 and L2) hyperpolarized at all wavelengths. L1 sensitivities peaked at 493 nm, near the G cone absorbance maximum. Modeled spectra suggest equally weighted inputs from both R and G cones and, in addition, a "hidden opponency" from blue cones. These were classified as R-/G-/(b+). L2 sensitivities were maximal at 563 nm near the R cone absorbance peak; modeled spectra were dominated by R cones, with lesser G cone contributions. B and UV cone signals were small or absent. These are R-/g-. Four chromatic (C-type) horizontal cells were either depolarized (+) or hyperpolarized (-) depending on stimulus wavelength. These types are biphasic (R+/G-/B-) with peak excitation at 467 nm, between G and B cone absorbance peaks, UV triphasic (r-/G+/U-) with peak excitation at 362 nm similar to UV cones, and blue triphasic (r-/G+/B-/u-) and blue tetraphasic (r-/G+/B-/u+), with peak excitation at 409 and 411 nm, respectively, similar to B cones. UV triphasic and blue tetraphasic horizontal cell spectral responses are unique and were not anticipated in previous models of distal color circuitry in cyprinids.

A spectral model for signal elements isolated from zebrafish photopic electroretinogram.

The zebrafish photopic electroretinogram (ERG) sums isolatable elements. In each element, red-, blue-, green-, and UV- (r, g, b, and u) cone signals combine in a way that reflects retinal organization. ERG responses to monochromatic stimuli of different wavelengths and irradiances were recorded on a white rod suppressing background using superfused eyecups. Onset elements were isolated with glutamatergic blockers and response subtractions. CNQX-blocked ionotropic (AMPA/kainate) glutamate receptors; l-AP4 or CPPG-blocked metabotropic (mGluR6) glutamate receptors; TBOA-blocked glutamate transporters; and l-aspartate inactivated all glutamatergic mechanisms. Seven elements emerged: photopic PIII, the l-aspartate-isolated cone response; b1, a CNQX-sensitive early b-wave element of inner retinal origin; PII, a photopic, CNQX-insensitive composite b-wave element from ON bipolar cells; PIIm, an l-AP4/CPPG-sensitive, CNQX-insensitive, metabotropic subelement of PII; PIInm, an l-AP4/CPPG/CNQX-insensitive nonmetabotropic subelement of PII; a1nm, a TBOA-sensitive, CNQX/l-AP4/CPPG-insensitive, nonmetabotropic, postphotoreceptor a-wave element; and a2, a CNQX-sensitive a-wave element linked to OFF bipolar cells. The first five elements were fit with a spectral model that demonstrates independence of cone-color pathways. From this, Vmax and half-saturation values (k) for the contributing r-, g-, b-, and u-cone signals were calculated. Two signal patterns emerged. For PIII or PIInm, the Vmax order was Vr > Vg >> Vb approximately Vu. For b1, PII, and PIIm, the Vmax order was Vr approximately Vb > Vg > Vu. In either pattern, u-cone amplitude (Vu) was smallest, but u-cone sensitivity (ku362) was greatest, some 10-30 times greater than r cone (kr570). The spectra of b1/PII/PIIm elements peaked near b- and u-cone absorbance maxima regardless of criteria, but the spectra of PIII/PIInm elements shifted from b- toward r-cone absorbance maxima as criterion levels increased. The greatest gains in Vmax relative to PIII occurred for the b- and u-cone signals in the b1/PII/PIIm b-wave elements. This suggests a high-gain prolific metabotropic circuitry for b- and u-cone bipolar cells.

Safety of high-dose corticosteroids for the treatment of autoimmune inner ear disease.

OBJECTIVE: To report the adverse effects associated with prolonged high-dose prednisone for the treatment of autoimmune inner ear disease (AIED). STUDY DESIGN: Prospective data collected as part of a multicenter, randomized, controlled trial for the treatment of corticosteroid-responsive AIED with methotrexate. SETTING: Tertiary referral centers. PATIENTS: One hundred sixteen patients with rapidly progressive, bilateral sensorineural hearing loss. INTERVENTION: All patients completed a 1-month course of prednisone (60 mg/d). In Phase 2, 67 patients with improvement in hearing underwent a monitored 18-week prednisone taper, resulting in 22 weeks of prednisone therapy at an average dose of 30 mg per day. Thirty-three patients were randomized to receive methotrexate in Phase 2. Thirty-four patients received prednisone and placebo. MAIN OUTCOME MEASURE: Adverse events (AE) in patients treated with prednisone only. RESULTS: Of 116 patients, 7 had to stop prednisone therapy during the 1-month challenge phase due to AE. Of 34 patients, 5 were unable to complete the full 22-week course of prednisone due to AE. The most common AE was hyperglycemia, which occurred in 17.6% of patients participating in Phase 2. Weight gain was also common, with a mean increase in body mass index of 1.6 kg/m2 (95% confidence interval, 0.77-2.3) during the 22-week steroid course. Patients entering Phase 2 were followed for a mean of 66 weeks. No fractures or osteonecrosis were reported. CONCLUSION: Although high-dose corticosteroids are associated with known serious side effects, prospective data in the literature are limited. The present study suggests that with appropriate patient selection, monitoring, and patient education, high-dose corticosteroids are a safe and effective treatment of AIED.

Electrophysiological evidence of GABAA and GABAC receptors on zebrafish retinal bipolar cells.

To refine inhibitory circuitry models for ON and OFF pathways in zebrafish retina, GABAergic properties of zebrafish bipolar cells were studied with two techniques: whole cell patch responses to GABA puffs in retinal slice, and voltage probe responses in isolated cells. Retinal slices documented predominantly axon terminal responses; isolated cells revealed mainly soma-dendritic responses. In the slice, GABA elicited a conductance increase, GABA responses were more robust at axon terminals than dendrites, and Erev varied with [Cl(-)]in. Axon terminals of ON- and OFF-type cells were similarly sensitive to GABA (30-40 pA peak current); axotomized cells were unresponsive. Bicuculline-sensitive, picrotoxin-sensitive, and picrotoxin-insensitive components were identified. Muscimol was as effective as GABA; baclofen was ineffective. Isolated bipolar cells were either intact or axotomized. Even in cells without an axon, GABA or muscimol (but not baclofen) hyperpolarized dendritic and somatic regions, suggesting significant distal expression. Median fluorescence change for GABA was -0.22 log units (approximately -16 mV); median half-amplitude dose was 0.4 microM. Reduced [Cl(-)]out blocked GABA responses. GABA hyperpolarized isolated ON-bipolar cells; OFF-cells were either unresponsive or depolarized. Hyperpolarizing GABA responses in isolated cells were bicuculline and TPMPA insensitive, but blocked or partially blocked by picrotoxin or zinc. In summary, axon terminals contain bicuculline-sensitive GABAA receptors and both picrotoxin-sensitive and insensitive GABAC receptors. Dendritic processes express zinc- and picrotoxin-sensitive GABAC receptors.

Transporter-mediated GABA responses in horizontal and bipolar cells of zebrafish retina.

GABA-mediated interactions between horizontal cells (HCs) and bipolar cells (BCs) transform signals within the image-processing circuitry of distal retina. To further understand this process, we have studied the GABA-driven membrane responses from isolated retinal neurons. Papain-dissociated retinal cells from adult zebrafish were exposed to GABAergic ligands while transmembrane potentials were monitored with a fluorescent voltage-sensitive dye (oxonol, DiBaC4(5)). In HCs hyperpolarizing, ionotropic GABA responses were almost never seen, nor were responses to baclofen or glycine. A GABA-induced depolarization followed by after hyperpolarization (dep/AHP) occurred in 38% of HCs. The median fluorescence increase (dep component) was 0.17 log units, about 22 mV. HC dep/AHP was not blocked by bicuculline or picrotoxin. Muscimol rarely evoked dep/AHP responses. In BCs picrotoxin sensitive, hyperpolarizing, ionotropic GABA and muscimol responses occurred in most cells. A picrotoxin insensitive dep/AHP response was seen in about 5% of BCs. The median fluorescence increase (dep component) was 0.18 log units, about 23 mV. Some BCs expressed both muscimol-induced hyperpolarizations and GABA-induced dep/AHP responses. For all cells, the pooled Hill fit to median dep amplitudes, in response to treatments with a GABA concentration series, gave an apparent k of 0.61 muM and an n of 1.1. The dep/AHP responses of all cells required both extracellular Na+ and Cl(-), as dep/AHP was blocked reversibly by Li+ substituted for Na+ and irreversibly by isethionate substituted for Cl(-). All cells with dep/AHP responses in zebrafish have the membrane physiology of neurons expressing GABA transporters. These cells likely accumulate GABA, a characteristic of GABAergic neurons. We suggest Na+ drives GABA into these cells, depolarizing the plasma membrane and triggering Na+, K+-dependent ATPase. The ATPase activity generates AHP. In addition to a GABA clearance function, these large-amplitude transporter responses may provide an outer plexiform layer GABA sensor mechanism.

Modulation of horizontal cell function by dopaminergic ligands in mammalian retina.

Light responses of rabbit horizontal cell somata (HC) to flickering light stimuli recorded with sharp electrodes consist of a distinctive flicker component superimposed on a sustained hyperpolarisation. Activation of dopamine D1/D5 receptors depolarises HC dark membrane potential and suppresses the flicker component of responses to photopic stimuli without affecting the sustained hyperpolarising response component. Waveforms of responses to scotopic stimuli are preserved. Similar response modulation was observed in depolarising cells of the inner retina, suggesting that activation of D1/D5 receptors of HC causes modification of cone signal transmission to higher order neurons. The impact of dopamine D1/D5 receptor activation on the function of HC in the light stimulated retina is discussed.

Fenretinide-induced neuronal differentiation of ARPE-19 human retinal pigment epithelial cells is associated with the differential expression of Hsp70, 14-3-3, pax-6, tubulin beta-III, NSE, and bag-1 proteins.

PURPOSE: We reported earlier that fenretinide can induce neuronal differentiation of ARPE-19 human retinal pigment epithelial cells in culture. The purpose of this study was to investigate the potential involvement of key proteins involved in gene transcription, signal transduction, cell cycle check point, differentiation, neuronal cell survival, and stress response in the neuronal differentiation of ARPE-19 cells by fenretinide. METHODS: Cells in culture were treated with 1.0 microM fenretinide. Cells were analyzed using antibodies against pax-6, neuronal specific enolase (NSE), tubulin beta-III, 14-3-3, bag-1, and Hsp-70 proteins using immunocytochemistry, western blot and ELISA methodologies. RESULTS: We found that pax-6 and NSE were both expressed in the control ARPE-19 cells. Fenretinide induced neuronal differentiation of ARPE-19 cells led to a decrease in pax-6 protein and an increase in tubulin beta-III protein expression after 5 days fenretinide treatment. There was a translocation of 14-3-3 from the cytoplasm to the nucleus, and an increase in nuclear expression of bag-1 after treatment. We also found a time-dependent increase in Hsp70 protein expression in ARPE-19 cells treated with fenretinide. D-407, another human retinal pigment epithelial cell line, but not either Y-79 or PC-12 cells, was also able to be induced into neuronal morphologies by fenretinide. CONCLUSIONS: The fenretinide-induced neuronal differentiation of ARPE-19 cells is associated with an increase in expression of the neuronal specific protein tubulin beta-III, and a decrease in expression of the progenitor cell marker pax-6. Neuronal differentiation of ARPE-19 cells is also associated with nuclear translocation of 14-3-3, a protein involved in signal transduction, cell cycle check point and cell growth, and an increase in expression of bag-1, a protein involved in neuronal cell survival and axon elongation. These results suggest that ARPE-19 cells could be a progenitor cell line that can be differentiated into neuronal cells when treated with factors such as fenretinide.