(from applicants' abstract) Given the rate at which evidence is amassing in support of astrocyte communication, it is important that the mechanisms underlying this communication be examined with the same rigor as was key to initially understanding the network behavior of neurons. Astrocytes are characterized in vivo as highly interconnected by gap junctions, and as having transmitter and receptor systems of unknown function. By reducing CNS complexity through established cell culture systems, both gap junctions and extracellular communication now have been found to serve roles in the propagation of calcium waves through networks of astrocytes. The passage of intracellular messengers through gap junctions has been recognized for some time as one mechanism for mediating calcium wave propagation. Recent results from this laboratory reveal the existence of an additional, and possibly dominant path: An extracellular message that is the focus of this proposal. We now know that astrocytes are capable of releasing an extracellular message which can be both necessary and sufficient for the sequential activation of neighboring astrocytes during the spreading calcium wave. The experiments in this proposal will identify the compound(s) serving as the extracellular messengers. They will examine the relative contribution of gap junction- versus extracellular message-mediated communication pathways for calcium wave propagation, and will test the hypothesis that these two paths represent parallel signaling routes. This work also will determine if the calcium transients that are the signature of the glial calcium wave have the function of evoking release of the extracellular message. Finally, the applicants will address a fundamental issue of the mechanisms of regenerative responses by individual astrocytes. They will ask whether an autocrine function, in which an astrocyte stimulates itself by the release of its own message, underlies the sequential activation of astrocytes along the course of the calcium wave. The questions addressed by this proposal are fundamental in themselves. The results of this proposal will therefore significantly enhance our ability both to interpret data already in the literature and affect the future experimental course of diverse research efforts.
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