The energy state of pancreatic beta cells : role in the control of insulin release
(1997)
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- Authors
- Supervisors
- Henquin, Jean-Claude
- Abstract
- Like most endocrine cells, pancreatic β cells undergo the influence of many extracellular messengers which interact with membrane receptors. However, the major regulators of insulin secretion are nutrients, in particular glucose, which have to be metabolized to exert their secretory action. This suggests that there exist particular second messengers and signal transduction pathways in β cells. Owing to the sensitivity of K+-ATP channels to ATP, adenine nucleotides have been proposed to serve as second messengers, but the ATP levels are two orders of magnitude higher than the Ki of the channel for ATP, and are not affected by glucose in the concentration range that stimulates insulin release. In this thesis, the possibility that changes in nucleotide concentrations in pancreatic β cells might regulate insulin secretion was carefully readdressed. <BR> We first characterized the distribution od adenine nucleotides between different subcellular pools, with a technique of permeabilization by the α-toxin from Staphylococcus Aureus. This permitted us to demonstrate that there exists within insulin secretory granules large pools of adenine nucleotides that is stable, insensitive to acute metabolic changes and has an ATP-ADP ratio close to 1. These pools, and in particular the pool of ADP, may mask the importance of the changes brought about by glucose in the metabolically active pools. <BR> To decrease the size of this background pool, the islets were partially degranulated by overnight culture In these islets, glucose induced a rapid and large increase in the ATP/ADP ratio. In the range of concentrations where it stimulates insulin release, glucose did not affect ATP levels but decreased ADP levels in a dose-dependant manner. This suggests that ADP may severe as a second messenger for the regulation of secretion, in particular at the level of the K+-ATP channels. <BR> In the presence of diazoxide and high K+, when glucose regulates secretion by a K+-ATP channel-dependent mechanism, a good correlation was also observed between the ATP/ ADP ratio and insulin release, suggesting a role for adenine nucleotides on a second site of regulation of insulin secretion. However, this second site of regulation of secretion by glucose is less sensitive to changes in the energy state of β cells than is the K+-ATP channel. <BR> We also tested the effects of glucose on guanine nucleotide levels in islets. Glucose induced a rapid increase in the CTP/GDP ratio that matched the increase in the ATP/ADP ratio. Under control conditions, insulin secretion was inversely correlated with the concentration of GDP, that might thus participate in the regulation of K+-ATP channels. Under conditions where the K+-ATP channel-independent effect of glucose was tested, insulin secretion was positively correlated with the GTP/GDP ratio, which suggests a role for guanine nucleotides also at this second site, possibly through regulation of the activity of G-proteins. <BR> In an attempts to identify which of the two purine nucleotide pools plays a regulatory role, we inhibited GMP synthesis with mycophenolic acid. The drug decreased GTP levels, impaired secretion, but also markedly affected the adenine nucleotide levels, suggesting that the two pools of nucleotides are so tightly interrelated that their respective functions cannot easily be selectively evaluated. <BR> Finally, we tested the relationchip between [Ca2+] and adenine nucleotide levels. When the glucose concentration was increased, the rise of the ATP/ADP ratio preceded that of [CA2+] and was not impaired in a Ca2+-free medium, indicating that Ca2+ is not necessary for the acceleration of cell metabolism by glucose. These observations reinforce the hypothesis that the rise in the ATP/ADP ratio triggers the entry of Ca2+ through modulation of the K+-ATP channels. Conversely, in the presence of a given glucose concentration, a stimulation of CA2+ entry was found to induce a rapid decrease in the ATP/ADP ratio whereas an inhibition of Ca2+ entry augmented the ratio. This suggests that oscillations of adenine nucleotides might take place during oscillations of [Ca2+]i. In an appendix, I propose a mathematical model for the generation of [Ca2+]I oscillations, base on a feed-back mechanism between [Ca2+]I and the ATP/ADP ratio. <BR> My contributions do not prove that adenine and guanine nucleotides serve as second messengers in β cells, but at least make plausible the hypothesis that the increase in the ATP/ADP ratio (or GTP/GDP ratio) is an important step for the regulation of insulin release by glucose
- Affiliations
Citations
de Timary, P. (1997). The energy state of pancreatic beta cells : role in the control of insulin release. https://hdl.handle.net/2078.5/111427