Abstract
Abstract: :
Purpose: Previous investigations have demonstrated the capability of the mammalian retina to synthesize melatonin. In addition, it has been shown that such synthesis is under the control of a circadian oscillator located within the retina. Several studies have shown that GABA modulates melatonin synthesis in the retina. An aim of the present investigation was to evaluate the possible role of GABA in the circadian mechanisms regulating melatonin synthesis Methods: Rat retinas were cultured at constant temperature (33ºC) in darkness for 3 days using a flow-through apparatus. The culture medium (199) was supplemented with 100 microM of GABA or GABA antagonists (saclofen and bicuculline). Perifusated samples were collected at 4 hour intervals, and melatonin levels were measured by radioimmnoassay. Results: Control retinas released melatonin with a clear circadian rhythm. Addition of 100 microM GABA to the medium did not affect the circadian rhythms of melatonin release; the same result was obtained culturing the retina with 100 microM of saclofen (GABAB antagonist). In contrast, the circadian rhythm of melatonin release/synthesis was abolished when the retinas were cultured with 100 microM of bicuculline (GABAA antagonist). Conclusion: Our data show that 100 microM of the GABAA antagonist bicuculline inhibited melatonin release in the rat retina, thus abolishing the circadian rhythm. Such effect could be a result of a direct action of GABA on the melatonin synthesizing cells (i.e., the cone photoreceptors) or an indirect action (i.e., throughout the increase in dopamine level which in turn inhibits melatonin synthesis). As the first step to understand the mechanism by which GABA acts we are now using laser captured microdissection and single cell RT-PCR to investigate if GABA receptors are present within the melatonin synthesizing cells.
Keywords: 349 circadian rhythms • 465 melatonin • 560 retinal culture