title: Phorbol ester impairs electrical excitation of rat pancreatic beta-cells through PKC-independent activation of KATP channels.
abstract: BACKGROUND
  Phorbol 12-myristate 13-acetate (PMA) is often used as an activating phorbol ester of protein kinase C (PKC) to investigate the roles of the kinase in cellular functions. Accumulating lines of evidence indicate that in addition to activating PKC, PMA also produces some regulatory effects in a PKC-independent manner. In this study, we investigated the non-PKC effects of PMA on electrical excitability of rat pancreatic beta-cells by using patch-clamp techniques.
  
  
  RESULTS
  In current-clamp recording, PMA (80 nM) reversibly inhibited 15 mM glucose-induced action potential spikes superimposed on a slow membrane depolarization and this inhibition can not be prevented by pre-treatment of the cell with a specific PKC inhibitor, bisindolylmaleimide (BIM, 1 microM). In the presence of a subthreshold concentration (5.5 mM) of glucose, PMA hyperpolarized beta-cells in a concentration-dependent manner (0.8-240 nM), even in the presence of BIM. Based on cell-attached single channel recordings, PMA increased ATP-sensitive K+ channel (KATP) activity. Based on inside-out patch-clamp recordings, PMA had little effect on KATP activity if no ATP was in the bath, while PMA restored KATP activity that was suppressed by 10 microM ATP in the bath. In voltage-clamp recording, PMA enhanced tolbutamide-sensitive membrane currents elicited by repetitive ramp pulses from -90 to -50 mV in a concentration-dependent manner, and this potentiation could not be prevented by pre-treatment of cell with BIM. 4alpha-phorbol 12,13-didecanoate (4alpha-PDD), a non-PKC-activating phorbol ester, mimicked the effect of PMA on both current-clamp and voltage-clamp recording configurations. With either 5.5 or 16.6 mM glucose in the extracellular solution, PMA (80 nM) increased insulin secretion from rat islets. However, in islets pretreated with BIM (1 microM), PMA did not increase, but rather reduced insulin secretion.
  
  
  CONCLUSIONS
  In rat pancreatic beta-cells, PMA modulates insulin secretion through a mixed mechanism: increases insulin secretion by activation of PKC, and meanwhile decrease insulin secretion by impairing beta-cell excitability in a PKC-independent manner. The enhancement of KATP activity by reducing sensitivity of KATP to ATP seems to underlie the PMA-induced impairment of beta-cells electrical excitation in response to glucose stimulation.
authors.affiliation: Department of Physiology, Hirosaki University School of Medicine, Hirosaki, 036-8562, Japan. ssuga@yahoo.com
authors.forename: S
authors.initials: S
authors.lastname: Suga
authors.forename: J
authors.initials: J
authors.lastname: Wu
authors.forename: Y
authors.initials: Y
authors.lastname: Ogawa
authors.forename: T
authors.initials: T
authors.lastname: Takeo
authors.forename: T
authors.initials: T
authors.lastname: Kanno
authors.forename: M
authors.initials: M
authors.lastname: Wakui
mesh_terms: D018528:ATP-Binding Cassette Transporters; D000818:Animals; D002273:Carcinogens / Q000494:pharmacology; D007515:Islets of Langerhans / Q000187:drug effects* / Q000378:metabolism; D054086:KATP Channels; D008297:Male; D010703:Phorbol Esters / Q000494:pharmacology; D015221:Potassium Channels / Q000378:metabolism*; D024661:Potassium Channels, Inwardly Rectifying; D011493:Protein Kinase C / Q000378:metabolism*; D051381:Rats; D017208:Rats, Wistar; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology*
chemical_list: D018528:ATP-Binding Cassette Transporters; D002273:Carcinogens; D054086:KATP Channels; D010703:Phorbol Esters; D015221:Potassium Channels; D024661:Potassium Channels, Inwardly Rectifying; C092520:uK-ATP-1 potassium channel; D011493:Protein Kinase C; D013755:Tetradecanoylphorbol Acetate