The relevance of depolarizing and non-depolarizing stimuli for the biphasic kinetics of insulin secretion.The role of the metabolic amplification.

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In this project we intend to gain further insight into the mechanisms which underlie the biphasic kinetics of insulin secretion, a feature which can be observed with human islets as well as with rodent islets. The depolarization-induced influx of Ca2+ is known to be necessary for the stimulated insulin secretion, however, the extent to which secretion is stimulated is determined by additional, currently incompletely understood signals. These signals are summarized as “amplifying pathway” or “metabolic amplification”. The stimulus-secretion-coupling in the pancreatic beta-cell has one specific feature: only those stimuli which are also nutrients for the beta-cell (“fuel secretagogues”) are able to induce a lasting increase of secretion. The fact that nutrient secretagogues have to be metabolized in order to stimulate insulin secretion has led to a bewildering multitude of potential signaling compounds in the amplifying pathway. Experiments with freshly isolated islets have led us to the hypothesis that the cataplerotic export of acetoacetate from the mitochondria to the cytosol and the ensuing increase of the cytosolic acetylCoA concentration could have a signaling function in the course of the metabolic amplification. How precisely cytosolic acetylCoA leads to increased secretion is an open question, but the reversible acetylation of certain proteins appears as a plausible mechanism. Such a reversible acetylation has been described for a number of proteins. Interestingly, regulators of the actin cytoskeleton can be found among these proteins. This leads to the following questions which we seek to answer in this project: 1. Can a signaling role of the export of acetoacetate and the resulting increase of cytosolic acetylCoA be demonstrated? 2. Is the function of the cytoskeleton affected by the reversible acetylation of certain regulatory proteins? 3. Finally, is there a relation between this acetylation and the mobility and fusion frequency of insulin granules?