Dichroism in Rods during Bleaching

Abstract

The long-lived intermediates in the rhodopsin bleaching sequence have been linked with the state of adaptation of isolated retinae (1, 2, 3, 4). Correlations of this type can only be made when the kinetics of the relevant dark reactions are established. Since spectrophotometric data for the isolated retina, on which kinetic analyses are normally based, are obtained using light passing axially through the rod outer segments (ROS), the concentrations of the intermediates at a given time after bleaching the retina can only be calculated if the orientation of each species with respect to the ROS axis is known. While the transverse orientation (i.e. perpendicular to the ROS long axis) of rhodopsin is well established (5, 6, 7, 8), the relative angles of the chromophores of the decay products are known less certainly. Denton’s measurements (6) in the near u. v. imply that metarhodopsin II (MII) is transversely orientated, and he deduced from polarized fluorescence data that retinol is aligned axially. In contrast, Wald et al. concluded that retinol lies in the same plane as rhodopsin (8). The present work seeks to establish the relative orientations of MII, metarhodopsin 465 (MIII), retinal and retinol in frog (Rana pipiens) and albino rat retinae. The technique used follows that of Denton (6) closely: the linear dichroism of rhodopsin and the long-lived products in the aligned rods on the edge of a folded retina were examined on a microspectrophotometer (MSP).