Publications (2)7.52 Total impact
-
Article: Quantification of retrograde axonal transport in the rat optic nerve by fluorogold spectrometry.
[show abstract] [hide abstract]
ABSTRACT: Disturbed axonal transport is an important pathogenic factor in many neurodegenerative diseases, such as glaucoma, an eye disease characterised by progressive atrophy of the optic nerve. Quantification of retrograde axonal transport in the optic nerve usually requires labour intensive histochemical techniques or expensive equipment for in vivo imaging. Here, we report on a robust alternative method using Fluorogold (FG) as tracer, which is spectrometrically quantified in retinal tissue lysate. To determine parameters reflecting the relative FG content of a sample FG was dissolved in retinal lysates at different concentrations and spectra were obtained. For validation in vivo FG was injected uni- or bilaterally into the superior colliculus (SC) of Sprague Dawley rats. The retinal lysate was analysed after 3, 5 and 7 days to determine the time course of FG accumulation in the retina (n = 15). In subsequent experiments axona transport was impaired by optic nerve crush (n = 3), laser-induced ocular hypertension (n = 5) or colchicine treatment to the SC (n = 10). Spectrometry at 370 nm excitation revealed two emission peaks at 430 and 610 nm. We devised a formula to calculate the relative FG content (c(FG)), from the emission spectrum. c(FG) is proportional to the real FG concentration as it corrects for variations of retinal protein concentration in the lysate. After SC injection, c(FG) monotonously increases with time (p = 0.002). Optic nerve axonal damage caused a significant decrease of c(FG) (crush p = 0.029; hypertension p = 0.025; colchicine p = 0.006). Lysates are amenable to subsequent protein analysis. Spectrometrical FG detection in retinal lysates allows for quantitative assessment of retrograde axonal transport using standard laboratory equipment. It is faster than histochemical techniques and may also complement morphological in vivo analyses.PLoS ONE 01/2012; 7(6):e38820. · 4.09 Impact Factor -
Article: Light scattering and wavefront aberrations in in vivo imaging of the rat eye: a comparison study.
[show abstract] [hide abstract]
ABSTRACT: In vivo imaging of the retina is becoming an increasingly important research method. General anesthesia rapidly compromises the corneal surface, which increases scattering. In addition, wavefront aberrations limit the maximum imaging resolution. Three common methods of stabilizing the air-cornea interface and reducing scattering are the use of a contact lens, a microscopy slide coverslip, or mineral oil. These methods have not yet been analyzed regarding their impact on scattering and wavefront aberrations. Nineteen eyes of 19 rats were analyzed with a custom-made Hartmann-Shack (HS) wavefront sensor. The amount of scattering was determined by analysis of the HS spot width, and the wavefront was reconstructed for the naked eye and each scattering-reducing method. Their effect on optical quality was determined by calculating the modulation transfer function (MTF). The three methods applied significantly reduced scattering but were differentially effective, with the coverslip performing the best and the mineral oil the worst. The root mean square (RMS) of the wavefront aberration, as well as the intereye variability of the RMS, was significantly smaller with the contact lens than with the coverslip. The MTF was best for the contact lens and worst for the coverslip, which was also illustrated by image simulations. The coverslip, contact lens, and mineral oil, when applied to the cornea, all reduced scattering. The best-performing method, the coverslip, increased wavefront aberrations. Overall, the contact lens had the best influence on image quality, and it appears to be the method of choice for high-resolution retinal imaging in rats.Investigative ophthalmology & visual science 05/2011; 52(7):4551-9. · 3.43 Impact Factor
