Live imaging of second messengers in developing retina

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The long-term objective is to delineate the cellular mechanisms by which spontaneous correlated neural activity is generated in the developing mammalian retina. Very early in brain development, before sensory experience is possible, both electrical and chemical activity is generated spontaneously throughout the immature nervous system. There is growing evidence that this early activity is critical for the appropriate development of neural circuits. Developing a detailed understanding of the organizing principles that govern the normal development of the human nervous system may make it possible to understand the origin of neurological birth defects. In addition, it will provide critical insights into devising strategies that allow the nervous system to rewire normal functioning neural circuits in response to developmental abnormalities such as amblyopia (lazy eye). The cellular basis of spontaneous activity in the retina has been studied primarily by electrophysiology and calcium imaging. Spontaneous retinal activity is characterized by depolarizations that occur with a period on the order of minutes. Indeed, we can reproduce this slow periodicity in dissociated retina neurons, indicating that the pacemaker underlying this periodicity may be cell autonomous. Here we propose to test the hypothesis that this slow periodicity is set by oscillations in the second messenger, cAMP. The goals of this proposal are two fold. First, we propose to implement in retinal neurons the use of two indicators - one sensitive to levels of the second-messenger cAMP levels, and the second sensitive to activity of protein kinase-A. Second, we will then use these indicators to determine whether spontaneous oscillations in cAMP underlie the periodicity observed in both the intact retina as well spontaneously active networks formed by dissociated retinal neurons.