No abstract

MATERIALS AND METHODSElectroretinography. Corneal electroretinograms (ERGs) were elicited by full-field 10-gs xenon photostrobe flashes and recorded at 0.1-1000 Hz by using methods and control values presented previously (6). Molecular Biology. Genomic DNA was isolated from peripheral blood samples. The Gly9OAsp mutation was found by sequencing the entire rhodopsin coding region, with target DNA generated by PCR amplification (7) of genomic DNA during 40 rounds of thermal cycling [94°C for 90 s, 54°C for 60 s, and 72°C for 180 s] followed by 72°C for 10 min. Exon 1 primers were 5'-AGCTCAGGCCTTCGCAGCAT-3' and 5'-GAGGGCTTTGGATAACATTG-3'. The codon 90 region was sequenced by the Sanger dideoxynucleotide chaintermination method (8) with primer 5'-ACGCAGCCCCTTC-GAGTAC-3'. The presence of the Gly9OAsp mutation was assayed in a population of normal subjects by allele-specific oligonucleotide hybridization with primers 5'-TGGT-GAAGCCACCTAGGAC-3' for Gly-90 and 5'-TGGTGA-AGTCACCTAGGAC-3' for Asp-90.Densitometry. The 530-nm test and 880-nm reference beams of a modified Florida retinal densitometer (9) were reflected from the human retina in vivo onto a photomultiplier tube. The 70 test field was centered on the 9.10 bleaching field. Measurements were made at 17.80 in the temporal retina of the left eye of the 38-year-old proband. A bleaching beam was either "white" or "monochromatic" from 10-nm interference filters (Baird-Atomic type B-1). Dark-corrected photon counts of test and reference beams were counted separately in 2-s intervals, and the ratio was averaged to give T, with To at the end of a long full bleach and Td after full dark recovery.(To -Td)/To = (1 -s)[1 -e-23(Am) [1] in which s is the fraction of dark-corrected test counts evoked by photons that have not passed twice through rhodopsin (i.e., stray light) and ,B is rhodopsin density (loge) expressed as a(Am)cl where a(Am) is the molecular extinction coefficient at the test wavelength Am, c is the concentration, and 1 is the optical path length through rhodopsin in the rod outer segment. Since s is not easily measured, it is common to assume that s = 0 and then to solve Eq. 1 for 2,. Converting to loglo gives the "two-way density" A, which is the frequently used parameter of retinal densitometry. At equilibrium, the kinetic equation of rhodopsin bleaching (10) is:Abbreviations: adRP, autosomal dominant retinitis pigmentosa; ERG, electroretinogram; sc-td, scotopic trolands; tvi, threshold versus retinal illuminance curve; Gly9OAsp, Gly-90 -> Asp. tTo whom reprint requests should be addressed at: W. K.

Reports of the upper limits of normal for lymph node size at abdominal computed tomography have varied from 6 to 20 mm. Establishment of an upper limit for node size by specific location, analogous to that which has been reported for mediastinal lymph nodes, was sought. Short-axis diameters of the lymph nodes were measured in 130 patients who were not likely to have enlarged abdominal lymph nodes. Seven locations were defined, and the largest nodal measurement for each was recorded. Histographic analysis and nonparametric statistical methods were used to determine threshold values for the maximum node size in each region. The upper limits of normal by location were as follows: retrocrural space, 6 mm; paracardiac, 8 mm; gastrohepatic ligament, 8 mm; upper paraaortic region, 9 mm; portacaval space, 10 mm; porta hepatis, 7 mm; and lower paraaortic region, 11 mm. Lower paraaortic lymph nodes larger than 11 mm by short-axis measurement are abnormal. In other locations, nodes smaller than 1 cm may be abnormal if the determined thresholds are exceeded.

SUMMARY1. Rhodopsin has been measured by Rushton's method of reflexion densitometry in a retinal region 180 temporal to the fovea, using a wavelength of measuring light (555 nm) so far into the long wave part of the spectrum that possible blue absorbing intermediates (e.g. transient orange) do not interfere.2. Rhodopsin was bleached by a strong light for 10 sec and then held steady by a weaker light. During a 10 see bleach, no regeneration occurs and the rate of bleaching is proportional to the quantum catch. The proportionality constant is about 10-7 (td sec)-'.3. From 2, the rate of photolysis at equilibrium produced by the steady light was calculated. Since conditions were at equilibrium, photolysis matched regeneration. It was found that the rate of regeneration was proportional to the amount of pigment still bleached. The proportionality constant was about 0-0025 sec-.4. It was found by several different methods that the constant in 3 is the same in the light or dark and hence regeneration occurs independently of bleaching.5. Therefore, the results from bleaching and regeneration experiments can be combined to give the general equation -400 dp/dt = Ip(10)444-(I where p is the fraction of rhodopsin, t is time in sec and I is the retinal illuminance.6. This equation describes the results of partial bleaching and regeneration experiments under a variety of different exposure intensities of moderately long (at least 10 min) exposure durations.7. The dark adaptation curve in a peripheral region of the rod monochromat's retina where there are few cones follows a simple exponential course over nearly 7 log1o units. Rhodopsin regeneration and log threshold for this region are described by the same curve with a time constant of about 400 sec. Each log unit fall in threshold is accompanied by 0 835 % increase in rhodopsin. This time constant is in agreement with Rushton's (1961) finding, but appreciably longer than that reported by Ripps & Weale (1969a).8. The Ripps & Weale result was, however, obtained by bleaching with a very short bright xenon flash (as they did). Under these conditions, blue absorbing intermediates) is (are) formed, the time constant of regeneration of rhodopsin is much faster than after long tungsten bleaches, and the kinetic equation is not valid.9. The general equation, together with the relation found in 7, successfully accounts for results previously published by others of the effect of duration and intensity of bleaching on the recovery of rod threshold in the dark, provided only that more than 5 % of the rhodopsin was bleached at the beginning of dark adaptation.

Foveal threshold elevation and red-green cone pigment regeneration have been studied in the dark after a wide range of bleaches in normal man with a view to probing the limits of the application of the Dowling-Rushton relation: i.e., the direct proportionality between log threshold elevation and fraction of unregenerated pigment. Cone pigment regeneration (and threshold recovery) is much faster after short bleaches than expected from the kinetics of a simple monomolecular reaction. Recovery is faster after a fixed (short) duration bleach the weaker it is. Except for the first 30 s after relatively weak bleaches and the entire recovery after a very brief (<0.001 s) saturating bright flash which bleaches a little more than 50 %, the results are accurately fit by the Dowling-Rushton relation over the entire range tested with only one arbitrary constant (the proportionality factor). Theory predicts too low threshold in comparison with what is obtained, for both of these exceptions It is a familiar fact that the recovery of visual sensitivity in the dark after significant bleaching proceeds only very slowly, and it was long presumed that this was because the visual pigment regenerated at a corresponding rate. However, experimental attempts to provide the justification for this presumption for rods went unrewarded until Dowling (1960), working with the electroretinogram of the rat, and Rushton (1961), measuring psychophysical thresholds on a human subject deficient in cones, found a linear relationship between the log threshold and the fraction of unregenerated rhodopsin. Their equation can be written log (Et/E 0 ) = a(l -p).