Role of the beta-cell cytoskeleton in metabolic and neurohormonal amplification of the two phases of insulin secretion
(2012)
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- Authors
- Supervisors
- Henquin, Jean-Claude
- Abstract
- (en) Glucose-induced insulin secretion is controlled by two pathways that depend on beta-cell metabolism. The triggering pathway starts by a rise in the cytosolic ATP/ADP ratio and closure of ATP-sensitive K+ channels in the plasma membrane, which leads to depolarization, opening of voltage-gated Ca2+ channels, influx of Ca2+ and an increase of cytosolic [Ca2+]c which induces exocytosis of insulin granules. In parallel a metabolic amplifying pathway augments the secretory response to Ca2+ via messengers and effectors that remain undetermined. Insulin secretion is also regulated by neural and hormonal signals which also mainly act as amplifying factors, i.e., they increase the secretory response triggered by glucose or another stimulus without affecting [Ca2+]c. I first investigated the involvement of beta-cell microfilaments and microtubules in metabolic amplification during the two phases of insulin secretion. I then evaluated whether beta-cell microfilaments are required for neurohormonal amplification and, simultaneously reevaluated the involvement of cAMP and PKC in metabolic amplification. Methods: Isolated mouse islets were cultured overnight before being treated with cytoskeleton-disrupting drugs and then perifused to measure insulin secretion and monitor [Ca2+]c changes. The proportions of free and polymerized actin and tubulin were determined in islet protein extracts. Metabolic amplification was mainly studied by comparing the Ca2+ and secretory responses to glucose and tolbutamide, a drug that mimics the triggering but not the amplifying effect of glucose. Results: I first established that metabolic amplification is involved in both phases of insulin secretion and not only in the second phase as suggested in most current models. I also found that both microfilament depolymerization and stabilization increased the secretory response, and that metabolic amplification persisted in the absence of functional microfilaments. My results then showed that disruption of microtubules does not affect insulin secretion whereas their stabilization slightly inhibits it partly because of a decrease in [Ca2+]c. Most importantly, functional microtubules proved unnecessary for metabolic amplification even during simultaneous disruption of microfilaments. In a third study, I established that amplification of insulin secretion by cAMP does not require actin microfilaments and that metabolic amplification is not mediated by cAMP. My fourth study showed that microfilaments are dispensable for amplification of insulin secretion by protein kinase C activation and I confirmed that the kinase is not involved in metabolic amplification. Conclusions: Metabolic amplification of insulin secretion is a rapid phenomenon involved in both phases of glucose-induced insulin secretion. It does not involve facilitation of insulin granule mobilization from reserve pools by the beta-cell cytoskeleton but affects a very distal step which I speculate to be acceleration of the priming of insulin granules possibly belonging to a highly Ca2+-sensitive pool. Neurohormonal amplification of insulin secretion is distinct from metabolic amplification (cAMP and PKC are not the mediators of the amplifying effect of glucose), but probably affects the same steps of insulin granule exocytosis independently of beta-cell microfilaments.
- Affiliations
Citations
Mourad, N. (2012). Role of the beta-cell cytoskeleton in metabolic and neurohormonal amplification of the two phases of insulin secretion. https://hdl.handle.net/2078.5/242024