Methods

Since 1976 (Stark & Zitzmann, 1976), a major thrust of the Stark lab research has concerned vitamin A deprivation and replacement. Vitamin A deprivation decreases rhodopsin in Drosophila rhabdomeres as inferred by spectroscopy, P-face particle density in freeze-fracture microscopy and electroretinographic (ERG) sensitivity (Harris et al, 1977). Carrot juice proved to be optimal in replacement experiments. Our first organized documentation of vitamin A replacement therapy was in 1991 (Sapp et al, 1991a): (1) cross-sectional area of rhabdomeres increased to asymptote at 2 days; (2) EM immunogold increased at 1 day and reached asymptote at 2 or 3 days; (3) absorbance difference recovered somewhat at 6 and 12 hr, most of the way at 1 day, and pretty much reached asymptote at 2 days. ERG sensitivity increased 100-fold within a few hours and reached asymptote at 3 days (Chen & Stark, 1992). Rhabdomeric P-face particle count showed an increase at 12.5 hr and reached the control (replete) count at 1 to two days (Stark & White, 1995); increases were also noted in the plasmalemma adjacent to the rhabdomeric membrane.

In summary, deprivation decreases rhodopsin without cell death and replacement synchronizes a massive de novo synthesis, export and deployment of rhodopsin witnessed by immunogold decoration of rough endoplasmic reticulum (Sapp et al, 1991b). Increased opsin gene transcription was suggested by promoter-reporter studies (Sun et al, 1993), northern blot analysis (Picking et al, 1996) and antibody staining {Shim, 2001 #1438}.

References

Chen DM, Stark WS (1992) Electrophysiological sensitivity of carotenoid deficient and replaced Drosophila. Vis Neurosci 9: 461-469

Harris WA, Ready DF, Lipson ED, Hudspeth AJ, Stark WS (1977) Vitamin A deprivation and Drosophila photopigments. Nature (Lond) 266: 648-650

Picking WL, Chen D-M, Lee RD, Vogt ME, Polizzi JL, Marietta RG, Stark WS (1996) Control of Drosophila opsin gene expression by carotenoids and retinoic acid: Northern and western analyses. Exp Eye Res 63: 493-500

Sapp RJ, Christianson JS, Maier L, Studer K, Stark WS (1991a) Carotenoid replacement therapy in Drosophila: recovery of membrane, opsin and visual pigment. Exp Eye Res 53: 73-79

Sapp RJ, Christianson JS, Stark WS (1991b) Turnover of membrane and opsin in visual receptors of normal and mutant Drosophila. J Neurocytol 20: 597-608

add Shim Mol Vis reference

Stark WS, White R, H. (1995) Carotenoid replacement in Drosophila: Freeze fracture electron microscopy. J Neurocytol sub: mitted

Stark WS, Zitzmann WG (1976) Isolation of adaptation mechanisms and photopigment spectra by vitamin A deprivation in Drosophila. J Comp Physiol 105: 15-27

Sun D, Chen DM, Harrelson A, Stark WS (1993) Increased expression of chloramphenicol acetyltransferase by carotenoid and retinoid replacement in Drosophila opsin promoter fusion stocks. Exp Eye Res 57: 177-187