Diminished Pupillary Light Reflex at High Irradiances in Melanopsin-Knockout Mice

Abstract

Abstract: : Purpose: In the mammalian retina, a small subset of retinal ganglion cells (RGCs) are intrinsically photosensitive. They express the opsin-like protein melanopsin and project to brain regions such as the olivary pretectal nucleus (OPN) and the suprachiasmatic nucleus (SCN), centers involved respectively in pupillary light reflex (PLR) and circadian photoentrainment, which are non-image-forming accessory visual functions. We want to assess quantitatively the function of this melanopsin-associated visual pathway by using the PLR as a simple and reliable assay. Methods: Previously, we have generated mice in which the melanopsin gene (mop) is replaced by a tau-LacZ coding sequence. We bred these animals to homozygosity (mop^-/-/tau-LacZ^+/+) and stained their isolated retinas and coronal brain sections with X-gal. Separately, RGCs innervating the SCN in mop^+/- and mop^-/- mice were labeled by retrograde transport of fluorescent beads, and recorded by whole-cell current clamp in isolated retinas to check for persistence of their intrinsic photosensitivity with 2mM Co²⁺ to block synaptic transmission. For measuring the consensual PLR, unanaesthetized mop^+/+, mop^+/-, mop^-/- and rd/rd cl ("rodless-coneless") mice dark-adapted for 1-3 hours were positioned with one eye subjected to illumination and the other eye imaged by a CCD camera in infrared light. Results: By X-gal labeling, RGCs that would normally express melanopsin were still present in mop^-/- mice, with normal number, morphology, and central projections to OPN and SCN. RGCs retrograde-labeled from the SCN in mop^+/- mice were still intrinsically photosensitive, but the corresponding RGCs from mop^-/- mice were not. The PLR of mop^+/- mice was like wild type. The PLR of mop^-/- mice was indistinguishable from wild type at low irradiances, but at high irradiances the reflex was incomplete. The PLR of rd/rd cl mice was highly light-insensitive at low irradiances, but reached completion at high irradiances. Conclusions: Melanopsin is absolutely required for the intrinsic photosensitivity of the RGCs in question. With respect to the PLR, the melanopsin-associated system and the classical rod/cone system appear to be complementary in function.

Keywords: circadian rhythms • pupillary reflex • ganglion cells