Light Activation of the Phosphoinositide Cycle in Intrinsically Photosensitive Chicken Retinal Ganglion Cells

Abstract

Purpose: In vertebrates, intrinsically photosensitive retinal ganglion cells (ipRGCs) acting as nonvisual photoreceptors transmit environmental illumination information to the brain, regulating diverse non–image-forming tasks. The phototransduction cascade in chicken ipRGCs has been shown to resemble that of rhabdomeric photoreceptors and involves phospholipase C (PLC) activation. The current work was an investigation of the participation of the phosphoinositide (PIP) cycle in this mechanism and of whether changes in activities of inositol 1,4,5-trisphosphate (IP3) and PIP kinase are triggered by light. Conclusions.: The results indicate for the first time that light stimulation of chicken RGC cultures activates the PIP cycle, causing an increase in intracellular levels of IP3, changes in levels of phosphatidic acid, PIP, and PIP2; and mobilization of Ca2+. Methods.: Primary cultures of Thy-1 immunopurified chicken embryonic RGCs were exposed to bright light pulses or kept in the dark, to assess intracellular Ca2+ mobilization by Fluo-3 AM fluorescence microscopy, IP3 levels, and enzymatic activities of diacylglycerol, phosphatidylinositol, and phosphatidylinositol phosphate kinases (DAGK, PIK, and PIPK, respectively), by radioactive assays. The presence of different melanopsins (Opn4m and Opn4x) and other photopigments was determined by RT-PCR and immunochemistry. Results.: Cultured RGCs expressing different nonvisual photopigments displayed a significant and rapid increase in IP3 levels (1.3-fold) and Ca2+ mobilization by light, which was reversed by administration of the PLC inhibitor U73122 (5 μM). Brief light pulses also caused a very rapid and transient activation of DAGK, PIK, and PIPK compared with that in the dark control.