Age-related changes in local water and protein content of human eye lenses measured by Raman microspectroscopy

The Raman microspectroscopic method was used to determine the local water and protein content in human lenses. In 18 lenses of varying age position-defined water/protein content measurements were carried out along the visual and the equatorial axis.\ud \ud A main characteristic of the human lens is its constant and relatively low protein content. In addition this constant nuclear value is reached within a short distance from the capsular surface. For statistical analysis of age-related changes the data points in individual lenses were piecewise linearized. (1) The mean nuclear water content was calculated from the data points in the inner 80% of the visual axis. (2) The steep drop in water content was linearized using a least-squares linear regression approach. The distance between lenticular surface and the intersection of the regression line with the line representing the nuclear mean was denominated as surface layer width.\ud \ud It proved that: (i) the mean nuclear water content significantly increased with age, (ii) the width of the surface layer was age independent in the anterior and posterior poles of the visual axis, and (iii) in the equatorial axis the surface layer width significantly decreased with age.\ud \ud Seven human lenses with small opaque spots were also investigated. The opaque spots proved to have a normal-for-site water content and some of them were flanked at their capsular side by a zone with a high-for-site water content.\ud \ud The correlation between protein content and refractive index and the observed decrease in nuclear protein content in aging human lenses can be taken as strong evidence that upon aging the refractive index of a major part of the human lens is reduced. The implications of this decrease is discussed with the respect to the problem known as the lens Paradox, i.e. the discrepancy between the theoretically expected age-related loss of far vision due to changes in lens curvature and axial position in the eye and the actually observed loss in near vision upon age

Local variation in absolute water content of human and rabbit eye lenses measured by Raman microspectroscopy

Raman spectra were obtained from fresh, fixed and sliced rabbit lenses and from human lens slices. For all lenses and lens slices the ratio R, defined as the Raman intensity at 3390 cm−1 divided by the Raman intensity at 2935 cm−1, was measured at different locations along the visual and equatorial axis. The ratios R were transformed to absolute water mass percentages by measuring solutions with known protein concentrations. It was shown that fixation and slicing have very little effect on the absolute water content of the lenses. The values obtained for the absolute water content are comparable to values given in literature. It was also shown that the water content in rabbit and human lenses rapidly decreases from the immediate anterior and posterior subsurface region to the deep superficial cortex and is relatively constant in the nucleus. Raman microspectroscopy appears to be a reliable method for the measurement of the absolute water content of small volumes on defined positions in the lens. This can be very useful when analyzing the possible relation between local variations in water content and the occurrence of opacities in the lens

Raman microspectroscopy of fixed rabbit and human lenses and lens slices: New potentialities

Raman spectroscopy is a non-invasive, non-destructive technique for the study of the macromolecular composition of tissues. Raman spectra were obtained from intact fresh and paraformaldehyde fixed rabbit lenses and from thin slices prepared from these lenses. In addition the Raman spectrum of an intact 82-yr-old human lens was compared with a slice of the same lens. It appeared that fixation with paraformaldehyde had only a minor qualitative effect on the Raman spectra and that Raman spectra of intact lenses and lens slices were comparable. It was also shown that in the slice of the old human lens the fluorescence, due to chromophores, could be reduced so that a reliable Raman spectrum could be obtained.\ud \ud The use of slices improves the accuracy of the position at which Raman spectra are recorded and fixation extends the time available for Raman analysis which is particularly important for the study of human lenses. Moreover, slicing enables Raman analysis of old human lenses, which up to now was thought to be impossible due to the overwhelming fluorescence of the chromophores present in these lenses

Square Arrays in Early Cortical Lens Opacities

A combined freeze-fracture and scanning electron microscopic study of early opaque spots in the aging human lens showed the absence of gap junctions and the presence of square arrays in the membranes of disturbed fibers and neighboring unaffected fibers. Square arrays, with membrane particles of 6-7 nm, are considered as rearranged gap junctions and/or intramembranous particles, with particle sizes between 8.5-9.S nm; they are a sign of electric and metabolic uncoupling. These ultrastructural observations lend support to the idea of an uncoupling mechanism in the aging human lens, conserving the transparency of unaffected parts of the lens, as postulated previously. Invest Ophthalmol Vis Sci 31:2476-2481, J990 As reemphasized by Bron and Brown, 1 most nuclear cataracts must be ascribed to increased light scattering due to increased amounts of high-molecular-weight proteins, and most cortical cataracts are the result of disturbance of the integrity of lens fibers. Cortical cataract is the most prevalent age-related type of cataract. 2 ' 3 Since cortical opacities in early stages tend to be stationary and as it is well established that lens fibers are extensively coupled by gap or communicating junctions

Retinal terminals in the goldfish optic tectum: Identification and characterization

Retinal terminal profiles in the goldfish optic tectum were identified electron microscopically after (1) labeling with horseradish peroxidase and (2) in the early stages of degeneration in short-term eye enucleates. All labeled terminals shared certain common morphological characteristics which were identical to those of a population of terminals in normal tecta. Terminals of this type disappeared 30 days after enucleation of the contralateral eye. Retinal terminal presynaptic profiles were characterized by (1) round and oval synaptic vesicles; (2) mitochondria with irregular, randomly oriented cristae, large intracristal spaces, dilated membrane spaces, and primarily light matrices; (3) a wide range in profile area, 0.06–6.82 Μm 2 ; (4) large numbers of synaptic vesicles per profile area 168± 33 synaptic vesicles per Μm 2 ; (5) asymmetric synapses; and (6) multiple synaptic contacts (1.46 ± 0.73 per terminal profile). The postsynaptic elements included both dendritic and, less commonly, pleomorphic vesicle-containing profiles. The majority of postsynaptic dendritic profiles were small (0.01–0.40 Μm 2 ). Serial synaptic contacts were occasionally seen. The combination of vesicular and mitochondrial morphology (1 and 2 above) was necessary and sufficient to establish the retinal origin of a terminal, but use of such criteria would underestimate the number of retinotectal terminals by omitting those which did not have a mitochondrion in the plane of section. The number of such terminals was calculated from independent measurements, and the total number of retinal terminal profiles per area of neuropil was estimated.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/50023/1/902260307_ftp.pd

The effects of denervation and stimulation upon synaptic ultrastructure

Quantitative studies of synaptic ultrastructure were made in the upper layers of cat cerebral cortex. Tissues were from intact cortex and from long-term (chronic) undercut cortex with or without electrical stimulation. The synaptic effects of chronic electrical stimulation of denervated cortex are most readily understood as growth and remodeling of synaptic elements. Associated with chronic stimulation were increases in: symmetric membrane contacts; areas of round and flat vesicle containing terminals; dendritic shaft contacts; and synaptic contact lengths. Even without stimulation there were indications of synaptic plasticity in denervated cortex; compared with intact cortex, synapses having symmetric membrane contacts showed an increase in bouton area and an increase in synaptic contacts on dendritic shafts. These data are consistent with the observations of others in which axonal terminal growth occurred after deafferentation. But it appears that chronic electrical stimulation in the adult nervous system promotes significantly more plasticity than occurs without stimulation. In a functional sense stimulation in the present experiments produced effective inhibition which did not occur with denervation alone. Thus the plasticity observed with stimulation had both structural and functional components.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/50005/1/901780107_ftp.pd