Measurements of membrane potential, transmembrane 45Ca fluxes, cytoplasmic free Ca2+ concentration and insulin release by transplantable rat insulinoma cells maintained in tissue culture

P. R. Flatt, H. Abrahamsson, P. Arkhammar, P. O. Berggren, P. Rorsman, S. K. Swanston-Flatt

Research output: Contribution to journalArticle

1 Citation (Scopus)

Abstract

Regulation of insulin release, membrane potential, transmembrane 45Ca fluxes and cytoplasmic free Ca2+ concentration, [Ca2+](i) was examined using suspension of transplantable NEDH rat insulinoma cells previously cultured for 2-3 days to eliminate necrotic tumour cells and counter prior hypoglycaemia. Insulinoma cells displayed a resting [Ca2+](i) of 94 ± 8 nM (n = 17) and released 104 ± 15 ng insulin 10-6 cells (n = 7) during 60 min incubation with uptake of 2.7 ± 0.2 nmol 45Ca 10-6 cells (n = 7). High concentrations of glucose did not affect membrane potential, transmembrane 45Ca fluxes [Ca2+](i) or insulin release by insulinoma cells. K+ at 25 mM depolarised the plasma membrane, induced a small increase in 45Ca efflux and increased [Ca2+](i) by 65%. This model action was not associated with demonstrable effects on 45Ca uptake and insulin release. The effect of 25 mM K+ on [Ca2+](i) was counteracted by D-600, but this blocker of voltage-activated Ca2+ channels and verapamil lacked effects on transmembrane 45Ca fluxes and insulin release. The Ca2+-calmodulin antagonist, trifluoroperazine, was also without effect on 45Ca fluxes and insulin release. Ca2+ ionophore ionomycin increased [Ca2+](i), whereas A23187 and X537A did not affect transmembrane 45Ca fluxes. Moreover, insulin release was independent of extracellular Ca2+ over the range 0-20.4 mM despite marked affects on transmembrane 45Ca fluxes and a greater than 4-fold change of [Ca2+](i). Dibutyryl cyclic AMP increased insulin release by 55% without affecting transmembrane 45Ca fluxes or [Ca2+](i). The phosphodiesterase inhibitor, theophylline, also enhanced insulin release by 10-36% with no change of 45Ca uptake. The effectiveness of theophylline was independent of extracellular Ca2+ over the range 0-10.2 mM. These results indicate that inappropriate Ca2+ regulation is a key pathogenic feature underlying the inappropriate insulin secretion of rat insulinoma cells.

Original languageEnglish
Pages (from-to)22-29
Number of pages8
JournalBritish Journal of Cancer
Volume58
Issue number1
StatePublished - Jan 1 1988
Externally publishedYes

Fingerprint

Insulinoma
Membrane Potentials
Insulin
Theophylline
Gallopamil
Trifluoperazine
Bucladesine
Ionomycin
Phosphodiesterase Inhibitors
Ionophores
Calcimycin
Calmodulin
Verapamil
Hypoglycemia
Cultured Cells
Suspensions
Cell Membrane
Glucose

ASJC Scopus subject areas

  • Cancer Research
  • Oncology

Cite this

Measurements of membrane potential, transmembrane 45Ca fluxes, cytoplasmic free Ca2+ concentration and insulin release by transplantable rat insulinoma cells maintained in tissue culture. / Flatt, P. R.; Abrahamsson, H.; Arkhammar, P.; Berggren, P. O.; Rorsman, P.; Swanston-Flatt, S. K.

In: British Journal of Cancer, Vol. 58, No. 1, 01.01.1988, p. 22-29.

Research output: Contribution to journalArticle

Flatt, P. R. ; Abrahamsson, H. ; Arkhammar, P. ; Berggren, P. O. ; Rorsman, P. ; Swanston-Flatt, S. K. / Measurements of membrane potential, transmembrane 45Ca fluxes, cytoplasmic free Ca2+ concentration and insulin release by transplantable rat insulinoma cells maintained in tissue culture. In: British Journal of Cancer. 1988 ; Vol. 58, No. 1. pp. 22-29.
@article{128ff59bcb7f478c93be286412764d07,
title = "Measurements of membrane potential, transmembrane 45Ca fluxes, cytoplasmic free Ca2+ concentration and insulin release by transplantable rat insulinoma cells maintained in tissue culture",
abstract = "Regulation of insulin release, membrane potential, transmembrane 45Ca fluxes and cytoplasmic free Ca2+ concentration, [Ca2+](i) was examined using suspension of transplantable NEDH rat insulinoma cells previously cultured for 2-3 days to eliminate necrotic tumour cells and counter prior hypoglycaemia. Insulinoma cells displayed a resting [Ca2+](i) of 94 ± 8 nM (n = 17) and released 104 ± 15 ng insulin 10-6 cells (n = 7) during 60 min incubation with uptake of 2.7 ± 0.2 nmol 45Ca 10-6 cells (n = 7). High concentrations of glucose did not affect membrane potential, transmembrane 45Ca fluxes [Ca2+](i) or insulin release by insulinoma cells. K+ at 25 mM depolarised the plasma membrane, induced a small increase in 45Ca efflux and increased [Ca2+](i) by 65{\%}. This model action was not associated with demonstrable effects on 45Ca uptake and insulin release. The effect of 25 mM K+ on [Ca2+](i) was counteracted by D-600, but this blocker of voltage-activated Ca2+ channels and verapamil lacked effects on transmembrane 45Ca fluxes and insulin release. The Ca2+-calmodulin antagonist, trifluoroperazine, was also without effect on 45Ca fluxes and insulin release. Ca2+ ionophore ionomycin increased [Ca2+](i), whereas A23187 and X537A did not affect transmembrane 45Ca fluxes. Moreover, insulin release was independent of extracellular Ca2+ over the range 0-20.4 mM despite marked affects on transmembrane 45Ca fluxes and a greater than 4-fold change of [Ca2+](i). Dibutyryl cyclic AMP increased insulin release by 55{\%} without affecting transmembrane 45Ca fluxes or [Ca2+](i). The phosphodiesterase inhibitor, theophylline, also enhanced insulin release by 10-36{\%} with no change of 45Ca uptake. The effectiveness of theophylline was independent of extracellular Ca2+ over the range 0-10.2 mM. These results indicate that inappropriate Ca2+ regulation is a key pathogenic feature underlying the inappropriate insulin secretion of rat insulinoma cells.",
author = "Flatt, {P. R.} and H. Abrahamsson and P. Arkhammar and Berggren, {P. O.} and P. Rorsman and Swanston-Flatt, {S. K.}",
year = "1988",
month = "1",
day = "1",
language = "English",
volume = "58",
pages = "22--29",
journal = "British Journal of Cancer",
issn = "0007-0920",
publisher = "Nature Publishing Group",
number = "1",

}

TY - JOUR

T1 - Measurements of membrane potential, transmembrane 45Ca fluxes, cytoplasmic free Ca2+ concentration and insulin release by transplantable rat insulinoma cells maintained in tissue culture

AU - Flatt, P. R.

AU - Abrahamsson, H.

AU - Arkhammar, P.

AU - Berggren, P. O.

AU - Rorsman, P.

AU - Swanston-Flatt, S. K.

PY - 1988/1/1

Y1 - 1988/1/1

N2 - Regulation of insulin release, membrane potential, transmembrane 45Ca fluxes and cytoplasmic free Ca2+ concentration, [Ca2+](i) was examined using suspension of transplantable NEDH rat insulinoma cells previously cultured for 2-3 days to eliminate necrotic tumour cells and counter prior hypoglycaemia. Insulinoma cells displayed a resting [Ca2+](i) of 94 ± 8 nM (n = 17) and released 104 ± 15 ng insulin 10-6 cells (n = 7) during 60 min incubation with uptake of 2.7 ± 0.2 nmol 45Ca 10-6 cells (n = 7). High concentrations of glucose did not affect membrane potential, transmembrane 45Ca fluxes [Ca2+](i) or insulin release by insulinoma cells. K+ at 25 mM depolarised the plasma membrane, induced a small increase in 45Ca efflux and increased [Ca2+](i) by 65%. This model action was not associated with demonstrable effects on 45Ca uptake and insulin release. The effect of 25 mM K+ on [Ca2+](i) was counteracted by D-600, but this blocker of voltage-activated Ca2+ channels and verapamil lacked effects on transmembrane 45Ca fluxes and insulin release. The Ca2+-calmodulin antagonist, trifluoroperazine, was also without effect on 45Ca fluxes and insulin release. Ca2+ ionophore ionomycin increased [Ca2+](i), whereas A23187 and X537A did not affect transmembrane 45Ca fluxes. Moreover, insulin release was independent of extracellular Ca2+ over the range 0-20.4 mM despite marked affects on transmembrane 45Ca fluxes and a greater than 4-fold change of [Ca2+](i). Dibutyryl cyclic AMP increased insulin release by 55% without affecting transmembrane 45Ca fluxes or [Ca2+](i). The phosphodiesterase inhibitor, theophylline, also enhanced insulin release by 10-36% with no change of 45Ca uptake. The effectiveness of theophylline was independent of extracellular Ca2+ over the range 0-10.2 mM. These results indicate that inappropriate Ca2+ regulation is a key pathogenic feature underlying the inappropriate insulin secretion of rat insulinoma cells.

AB - Regulation of insulin release, membrane potential, transmembrane 45Ca fluxes and cytoplasmic free Ca2+ concentration, [Ca2+](i) was examined using suspension of transplantable NEDH rat insulinoma cells previously cultured for 2-3 days to eliminate necrotic tumour cells and counter prior hypoglycaemia. Insulinoma cells displayed a resting [Ca2+](i) of 94 ± 8 nM (n = 17) and released 104 ± 15 ng insulin 10-6 cells (n = 7) during 60 min incubation with uptake of 2.7 ± 0.2 nmol 45Ca 10-6 cells (n = 7). High concentrations of glucose did not affect membrane potential, transmembrane 45Ca fluxes [Ca2+](i) or insulin release by insulinoma cells. K+ at 25 mM depolarised the plasma membrane, induced a small increase in 45Ca efflux and increased [Ca2+](i) by 65%. This model action was not associated with demonstrable effects on 45Ca uptake and insulin release. The effect of 25 mM K+ on [Ca2+](i) was counteracted by D-600, but this blocker of voltage-activated Ca2+ channels and verapamil lacked effects on transmembrane 45Ca fluxes and insulin release. The Ca2+-calmodulin antagonist, trifluoroperazine, was also without effect on 45Ca fluxes and insulin release. Ca2+ ionophore ionomycin increased [Ca2+](i), whereas A23187 and X537A did not affect transmembrane 45Ca fluxes. Moreover, insulin release was independent of extracellular Ca2+ over the range 0-20.4 mM despite marked affects on transmembrane 45Ca fluxes and a greater than 4-fold change of [Ca2+](i). Dibutyryl cyclic AMP increased insulin release by 55% without affecting transmembrane 45Ca fluxes or [Ca2+](i). The phosphodiesterase inhibitor, theophylline, also enhanced insulin release by 10-36% with no change of 45Ca uptake. The effectiveness of theophylline was independent of extracellular Ca2+ over the range 0-10.2 mM. These results indicate that inappropriate Ca2+ regulation is a key pathogenic feature underlying the inappropriate insulin secretion of rat insulinoma cells.

UR - http://www.scopus.com/inward/record.url?scp=0023687231&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=0023687231&partnerID=8YFLogxK

M3 - Article

C2 - 2844219

AN - SCOPUS:0023687231

VL - 58

SP - 22

EP - 29

JO - British Journal of Cancer

JF - British Journal of Cancer

SN - 0007-0920

IS - 1

ER -