Microspectrophotometry ofDrosophila visual pigments: Determinations of conversion efficiency in R1–6 receptors

Summary

We used microspectrophotometry, together with some associated electrophysiological measurements, to characterize the photopigments of R1–6 receptors in living, white-eyedDrosophila. Measurements were from the deep pseudopupil, an optical visualization of photopigment-containing rhabdomeres. Transmission changes associated with photointerconversion of rhodopsin (R480) and its stable metarhodopsin (M570) are easily seen (Fig. 1). Such transmission changes were measured to produce a difference specrum (Fig. 2); the isosbestic wavelength is about 502 nm. A photoequilibrium spectrum (Fig. 3), analyzed together with sensitivity data (Fig. 4a), was used to determine the fraction of M570 in photoequilibria established with various monochromatic wavelengths. From this, the quantum efficiency for M570 to R480 conversion relative to R480 to M570 efficiency was determined to be about 0.71. No dark regeneration of M570 to R480 occurred within a period of 60min (Fig. 4a, b).

The extent of conversion related to incident energy was estimated as a function of wavelength (Figs. 5 and 6). These experiments yielded the photosensitivity spectrum of the visual pigment (Fig. 6). Assuming that the absorption spectrum of rhodopsin (R480) is the same as the sensitivity spectrum as determined by electrophysiology, this photosensitivity spectrum was used to derive the spectrum of metarhodopsin (M570) (Fig. 7). The maximal extinction of M570 is about 1.43 times the maximal extinction of R480. The spectra of both states of the photopigment fit the Ebrey-Honig nomogram in the long wavelength band. Both of the photosensitivity spectra, measured in the living eye, associated with R480 and M570 respectively have a major UV maximum.