CNG-modulin, The Cone Specific Modulator Of CNG Channel Activity, Is Required For The Recovery Of Flash Sensitivity Under Continuing Illumination Characteristic Of Cone Photoreceptors

Abstract

Purpose: : The ligand sensitivity of cGMP-gated ion channels in cone photoreceptors changes as a function of Ca²⁺ and this effect is mediated by CNG-modulin, a newly discovered Ca²⁺ binding protein expressed in cones, but not rods. Sensitivity modulation renders channel activity dependent on both cGMP and Ca²⁺ and helps control the light-sensitivity, time course and stability of the photoresponse. Suppression of CNG-modulin expression in zebrafish (ZF) cones is used to test one specific function possibly associated with CNG-modulin: the slow recovery of sensitivity in the continuing presence of steady light.

Methods: : A morpholino antisense oligonucleotide complementary to the amino terminus of ZF CNG-modulin was injected into ZF embryos. 5-6 days later the expression of CNG-modulin in cones was assayed with immunohistochemistry and the cone electrical photoresponse was measured from the a-wave of ergs recorded in intact larvae incubated in APB (2-amino-4-phosphonobutyric acid), a well-established blocker of non-photoreceptor erg components.

Results: : 5-6 days post fertilization the retina of wild type ZF larva have mature, normally functioning cone photoreceptors, but not rods. The isolated erg a-wave in the wild type larva shows that under continuous illumination, the cone photoresponse to test flashes superimposed on light steps is initially small, but recovers in amplitude with an exponential time course of 2-3 sec time constant. Anti CNG-modulin morpholino-injected larva, in contrast, responds to the same test flashes with the same initial small response, but the response amplitude does not recover in amplitude at all over the following 15 sec. Immunohistochemistry demonstrates that CNG-modulin is expressed in the cones of wild type larvae, but not in those of morpholino-injected ones.

Conclusions: : Slow sensitivity recovery in the continuous presence of light characteristic of normal cones does not occur in cones in which the expression of CNG-modulin is suppressed by gene knock-down. This indicates that sensitivity recovery depends on the Ca²⁺-dependent modulation of CNG ion channel activity mediated by CNG-modulin.

Keywords: photoreceptors • ion channels • calcium