Lessons from photoreceptors: turning off g-protein signaling in living cells

Abstract

Phototransduction in retinal rods is one of the most extensively studied G-protein signaling systems. In recent years, our understanding of the biochemical steps that regulate the deactivation of the rod's response to light has greatly improved. Here, we summarize recent advances and highlight some of the remaining puzzles in this model signaling system.

FIGURE 1. The rod photoreceptor and the disc-associated proteins that mediate the response to light

A: rod photoreceptors are highly compartmentalized cells, with most organelles retained in the inner segment (IS) and synaptic ending (SE). The specialized outer segment (OS) is filled with a stack of intracellular membranous discs that house the transduction machinery. B: schematic of a portion of three disc membranes and plasma membrane shown in A, illustrating activation and deactivation reactions.

FIGURE 2. Kinetic schemes of phototransduction deactivation

A: standard schemes for R* and G*-E* deactivation, each assuming a single rate constant (k_R or k_E, respectively) for the shut-off reactions. B: expanded schemes for the deactivation events boxed in A, which include rate constants for kinase (k_k) and arrestin (k_a) actions known to be essential for R* deactivation and the forward (k_f), reverse (k_b), and catalytic (k_cat) rate constants needed to describe the RGS9-catalyzed GTP hydrolysis needed for G*-E* deactivation.

FIGURE 3. Kinetics of responses recorded from mouse rods using suction electrodes closely match the kinetics of in vivo electroretinographic (ERG) recordings from mice

A: rod photoresponse extracted from paired-flash ERG experiments of Hetling and Pepperberg (symbols; Ref. ; Figure 5B); flash produced a response of ~55% the saturating a-wave amplitude corneal ERG a-waves (symbol). The black trace is a population average half-maximal response from 15 c57Bl/6 mouse rods recorded with suction electrodes; the flash strength was 48 photons/µm². Red curves are exponential functions, both with a 202-ms time constant. B: paired-flash mouse a-wave data of Lyubarsky and Pugh (1996) for a series of flashes of increasing strength that completely suppressed the a-wave (symbols) are compared with population average suction electrode recordings (smooth traces) from 15 mouse rods made in bicarbonate-buffered Locke’s solution at 37°C to flashes that ranged from 580 to 25,200 pho-tons/µm² by factors of 2. The average maximal response amplitude for the suction electrode recordings in A and B was 16.5 pA; the average effective collecting area was 0.50 µm².

FIGURE 4. Dark-adapted responses of a wild-type mouse rod

A: population average response family from a Sv129 mouse rod. Brief (10 ms) flashes were delivered at t = 0 s and ranged in strength from 7.78 to 85,600 photons/µm² by factors of 2–4. B: Pepperberg plot of wild-type mouse rods. The time that a bright flash response remained in saturation (T_sat) is plotted as a function of the natural logarithm of the flash strength in photoisomerizations per flash (R*). The slopes of the fitted lines, 0.26 and 0.80 s, give the dominant time constants of recovery over the respective range of flash strengths.