Enhanced stimulus-secretion coupling in polyamine-depleted rat insulinoma cells. An effect involving increased cytoplasmic Ca2+, inositol phosphate generation, and phorbol ester sensitivity

A. Sjoholm, P. Arkhammar, N. Welsh, K. Bokvist, P. Rorsman, A. Hallberg, T. Nilsson, M. Welsh, P. O. Berggren

Research output: Contribution to journalArticle

32 Scopus citations

Abstract

To extend previous observations on the role of polyamines in insulin production, metabolism, and replication of insulin-secreting pancreatic β cells, we have studied the role of polyamines in the regulation of the stimulus-secretion coupling of clonal rat insulinoma cells (RINm5F). For this purpose, RINm5F cells were partially depleted in their polyamine contents by use of the specific ornithine decarboxylase inhibitor difluoromethylornithine (DFMO), which led to an increase in cellular insulin and ATP contents. Analysis of different parts of the signal transduction pathway revealed that insulin secretion and the increase in cytoplasmic free Ca2+ concentration ([Ca2+]) after K+-induced depolarization were markedly enhanced in DFMO-treated cells. These effects were paralleled by increased voltage-activated Ca2+ currents, as judged by whole-cell patch-clamp analysis, probably reflecting increased channel activity rather than elevated number of channels per cell. DFMO treatment also rendered phospholipase C in these cells more sensitive to the muscarinic receptor agonist carbamylcholine, as evidenced by enhanced generation of inositol phosphates, increase in [Ca2+]i and insulin secretion, despite an unaltered ligand binding to muscarinic receptors and lack of effect on protein kinase C activity. In addition, the tumor promoter 12-O-tetradecanoylphorbol 13-acetate, at concentrations suggested to be specific for protein kinase C activation, evoked an increased insulin output in polyamine-deprived cells compared to control cells. The stimulatory effects of glucose or the cyclic AMP raising agent theophylline on insulin release were not increased by DFMO treatment. In spite of increased binding of sulfonylurea in DFMO-treated cells, there was no secretory response or altered increase in [Ca2+]i in response to the drug in these cells. It is concluded that partial polyamine depletion sensitizes the stimulus-secretion coupling at multiple levels in the insulinoma cells, including increased voltage-dependent Ca2+ influx and enhanced responsiveness to activators of phospholipase C and protein kinase C. In their entirety, our present results indicate that the behavior of the stimulus-secretion coupling of polyamine-depleted RINm5F insulinoma cells changes towards that of native β cells, thus improving the use-7ulness of this cell line for studies of β cell insulin secretion.

Original languageEnglish (US)
Pages (from-to)1910-1917
Number of pages8
JournalJournal of Clinical Investigation
Volume92
Issue number4
DOIs
StatePublished - 1993

Keywords

  • Ca channels
  • Inositol trisphosphate ·
  • Insulin secre-tion ·
  • Polyamides ·
  • Protein kinase c ·

ASJC Scopus subject areas

  • Medicine(all)

Fingerprint Dive into the research topics of 'Enhanced stimulus-secretion coupling in polyamine-depleted rat insulinoma cells. An effect involving increased cytoplasmic Ca<sup>2+</sup>, inositol phosphate generation, and phorbol ester sensitivity'. Together they form a unique fingerprint.

  • Cite this