an autonomous circadian clock in the inner mouse retina regulated by dopamine and gaba自主生物钟的老鼠视网膜由多巴胺和gaba监管.pdfVIP

PLoS BIOLOGY An Autonomous Circadian Clock in the Inner Mouse Retina Regulated by Dopamine and GABA * Guo-Xiang Ruan, Gregg C. Allen, Shin Yamazaki, Douglas G. McMahon Department of Biological Sciences, Vanderbilt University, Nashville, Tennessee, United States of America The influence of the mammalian retinal circadian clock on retinal physiology and function is widely recognized, yet the cellular elements and neural regulation of retinal circadian pacemaking remain unclear due to the challenge of long- term culture of adult mammalian retina and the lack of an ideal experimental measure of the retinal circadian clock. In the current study, we developed a protocol for long-term culture of intact mouse retinas, which allows retinal circadian rhythms to be monitored in real time as luminescence rhythms from a PERIOD2::LUCIFERASE (PER2::LUC) clock gene reporter. With this in vitro assay, we studied the characteristics and location within the retina of circadian PER2::LUC rhythms, the influence of major retinal neurotransmitters, and the resetting of the retinal circadian clock by light. Retinal PER2::LUC rhythms were routinely measured from whole-mount retinal explants for 10 d and for up to 30 d. Imaging of vertical retinal slices demonstrated that the rhythmic luminescence signals were concentrated in the inner nuclear layer. Interruption of cell communication via the major neurotransmitter systems of photoreceptors and ganglion cells (melatonin and glutamate) and the inner nuclear layer (dopamine, acetylcholine, GABA, glycine, and glutamate) did not disrupt generation of retinal circadian PER2::LUC rhythms, nor did interruption of intercellular communication throu