Mechanisms Regulating Variability of the Single Photon Responses of Mammalian Rod Photoreceptors

{{Summary
 * title=Mechanisms Regulating Variability of the Single Photon Responses of Mammalian Rod Photoreceptors
 * authors=Greg D. Field, Fred Rieke
 * url=http://www.cns.nyu.edu/events/vjclub/archive/field2002.pdf
 * tags=single photon response, photoreceptors, vision
 * summary=Rod photoreceptors have good single-photon responses, but variability in that response limits the accuracy and timing of photon absorption. This experiment studies how much fluctuation takes place, and what mechanisms are involved.  Rods of mammals and toads are tested to and their fluctuations compared to those of single molecules, to inform a model of single photon response.

Goals and Methods
A weak fixed-strength flash was repeated to primate and guinea pig rods. They both generate quantized responses to such stimulus. Flash intensity was chosen to stimulate an average of 0.5 photoisomerization of Rhodopsin. The dominant source of variability trial to trial in the response was expected to be Poisson fluctuation in the base current and the photon response.

Single photon response was separated from failures (no response) and multiphoton response (easy to identify as a multiple of the single-photon resopnse - either 2x or 3x). This was made easier by the weakness of the flash.

Singles and failures were isolated across 3-4 flash strengths from each tested cell, to analyze contamination: as measured by the lack of dependence of isolated singles. Rods with little observed contamination were used for the rest of the experiment.

Mechanisms were identified that might limit the single photon response:
 * 1) Local depletion of cGMP
 * 2) Depletion of the available transducin or PDE
 * 3) Feedback loops in rhodopsin activity
 * 4) Multistep shutoff

The first would imply different results from uniform and localized illumination, which was not observed. The second, third, and fourth were all parameterized and optimized to try to fit the observed mean data.

Results and Analysis
The fluctuations of rod responses to single photons are 3-4x smaller than other biological signals produced by single molecules. This data is used to constrain models for how that photon response is controlled.

Simple feedback of rhodopsin activity or transduction cascade saturation alone could not produce the observed results - they were perhaps too simple but also had the wrong relationwhip between their mean and variance. . The best model for single photon response identified was a multistep process involving some combination of rhodopsin shutoff and saturation.

This model was then applied to the shape of the response curve under the load of a buffer (BAPTA). Without changing the parameters of the model, it continued to fit this new scenario well. Combinations of most models failed to improve on the multistep model, but a saturation + multistep model could potentially reduce the number of steps.

The authors however suggest that a multistep shutoff alone is the most likely candidate process. In this case molecular constraints on rhodopsin activation and reaction rates would include the need to have reverse rate constants ~20x smaller than the forward constant for a 10-step process. That would require a large free energy difference between active and inactive rhodopsin - in this case on the order of 20kcal/mol.

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 * relevance=This provides baseline data on the fluctuations of mammalian rods to photons, and suggests a multistep model for rhodopsin shutoff and saturation that could account for it.
 * journal=Neuron
 * pub_date=2002/08/15
 * doi=10.1016/S0896-6273(02)00822-X
 * subject=Neuroscience