J Physiol 586.22 (2008) p 5291 5291

PERSPECT IVES

Glucose-induced insulinsecretion: is the small G-proteinRab27A the mediator of the KATP

channel-independent effect?

Susanne UllrichUniversity of Tubingen, Departmentof Internal Medicine, Divisionof Endo-crinology, Diabetology, Angiology,Nephrology and Clinical Chemistry,Tubingen, Germany

Email: susanne.ullrich@med.uni-tuebingen.de

From yeast to neurones, living cellscommunicate with their environment bysecretion of substances through regulatedexocytosis. The substances are stored insecretory granules within the cytosol. As thecells, their needs and function are extremelydivergent, synthesis, storage and secretionof cellular products are adapted.

Although the regulation of exocytosisis cell specific the secretory machineriesare amazingly similar. Whether in yeast,neurone or insulin-secreting cells SNAREproteins enable the fusion of the twomembranes, the vesicular and plasmamembrane and the release of cell-madematerial. SNARE protein interaction, i.e.fusion, is largely regulated by Ca2+. SmallG proteins comprise another protein familythat is evolutionary highly conserved.Distinct members of this family regulatethe formation, the directed trafficking andthe fusion of vesicles. Rab3A and Rab27Aare two small G proteins identified tobe involved in insulin release (Regazziet al. 1992; Olszewski et al. 1994; Yiet al. 2002). Another vesicular protein,granuphilin that mediates granule dockingto the plasma membrane is an effector ofRab27A. Kasai et al. (2005) demonstratedthat rab27A-deficient ashen mice exhibitglucose intolerance due to insufficientglucose-induced insulin secretion whilegranuphilin-deficient insulin secreting cellsrelease a higher amount of insulin. Theseobservations are puzzling and suggestthat granuphilin inhibits while Rab27Apromotes granule fusion.

Measurement of membrane capacitanceusing the patch clamp method allows theon-line observation of exo- and endo-cytotic events in cells under voltage clamp

and the analysis of kinetic changes ofplasma membrane surface area. Briefdepolarizing voltage pulses are applied tomimic glucose-induced action potentialsthat result in Ca2+ influx throughvoltage-dependent Ca2+ channels. Theimmediate quantal increase in capacitancehas been proposed to represent the readilyreleasable pool (RRP) of already dockedinsulin-containing vesicles (Olofsson et al.2002). With a rise in glucose concentration,the increase in capacitance is enlarged dueto a recruitment of additional granules tothe RRP. This effect of glucose is mimickedby larger and repetitive depolarizing pulses.Shorter and smaller depolarization resultsin smaller increases in capacitance fromthe immediate releasable pool (IRP) ofgranules. It seems logical that the RRPrepresents the first, rapid phase of insulinsecretion after glucose stimulation whilethe second slow phase of secretion whenglucose remains high may mainly resultfrom granules recruited to the plasmamembrane from the reserve pool.

The study published in this issue of TheJournal of Physiology by Merrins & Stuenkel(2008) used the approach of capacitancemeasurements to examine the effect ofrab27A deficiency in ashen islet cells on IRPand RRP mobilization and refilling. Via apulse protocol of five short (50 ms) andeight long (500 ms) depolarizing pulses,the size of IRP and RRP, respectively,was analysed. The repetition of the pulseprotocol allowed the analysis of the refillingof the pools. The capacitance changesto the first pulse trail were not differentbetween wild type and ashen mouseislet cells while the answer to the secondpulse trail was reduced in ashen islet cells.This observation confirms convincingly thatdocking, i.e. IRP and RRP, is not changed byRab27A deficiency but refilling the pools.This refilling deficiency was antagonized bycAMP, an effect blocked by the inhibitionof protein kinase A, indicating that cAMPfacilitates exocytosis independent ofRab27A expression. Increasing the glucoseconcentration incompletely restoredrefilling of RRP in ashen islet cells.Interestingly, the deficiency of refilling isthus specific for glucose.

Total internal reflection microscopy(TRIFM), a method that allows the analysisof granule movement and fusion in livingcells, was used by Nagamatsu and colleagues

(Nagamatsu, 2006). They confirmed thatafter glucose stimulation, mainly dockedgranules are released immediately followedby recruited granules. Kasai et al. (2008)present a detailed analysis of the provenanceof granules during K+- and glucose-inducedinsulin release. They found that K+ initiatedsecretion within 5 s and mainly dockedgranules (80%) underwent fusion. Incontrast, glucose-induced exocytosis startedafter 20 s and only 30% of granulesoriginated from the docked pool but 70%from the reserve pool. In ashen mice,granule fusion in islet cells induced byglucose but not by K+ was reduced mainlydue to reduced recruitment from the reservepool.

These observations support a new conceptthat IRP and RRP are granule pools withvariable Ca2+ sensitivities and are dockedto and blocked at the plasma membranevia granuphilin. These granules becomemore sensitive to Ca2+-mediated fusionwhen granuphilin-dependent blocking isameliorated by Rab27A. Glucose exerts adual effect: it stimulates docked granulesthrough Ca2+ influx as does K+ and via anunknown mechanism it stimulates Rab27Athat activates docked granules as well asrecruits granules to the plasma membrane.In this concept Rab27A may mediate theKATP channel-independent effect of glucoseon insulin secretion.

References

Kasai K, Fujita T, Gomi H & Izumi T (2008).Traffic 9, 1191–1203.

Kasai K, Ohara-Imaizumi M, Takahashi N,Mizutani S, Zhao S, Kikuta T, Kasai H,Nagamatsu S, Gomi H & Izumi T (2005). JClin Invest 115, 388–396.

Merrins MJ & Stuenkel EL (2008). J Physiol 586,5367–5381.

Nagamatsu S (2006). Endocr J 53, 433–440.Olofsson CS, Gopel SO, Barg S, Galvanovskis J,

Ma X, Salehi A, Rorsman P & Eliasson L(2002). Pflugers Arch 444, 43–51.

Olszewski S, Deeney JT, Schuppin GT, WilliamsKP, Corkey BE & Rhodes CJ (1994). J BiolChem 269, 27987–27991.

Regazzi R, Vallar L, Ullrich S, Ravazzola M,Kikuchi A, Takai Y & Wollheim CB (1992).Eur J Biochem 208, 729–737.

Yi Z, Yokota H, Torii S, Aoki T, Hosaka M, ZhaoS, Takata K, Takeuchi T & Izumi T (2002). MolCell Biol 22, 1858–1867.

C© 2008 The Author. Journal compilation C© 2008 The Physiological Society DOI: 10.1113/jphysiol.2008.164095