Effect of buffer systems and pH(i) on the measurement of [Ca2+](i) with fura 2

M. B. Ganz, J. Rasmussen, Wendy B Bollag, H. Rasmussen

Research output: Contribution to journalArticle

36 Citations (Scopus)

Abstract

The fluorescent probe, fura 2, is widely used to measure agonist-induced changes in intracellular calcium concentration ([Ca2+](i)) in cultured cells. However, in many instances, the results obtained in the same cell type have differed from one study to the next. The possibility that such differences might be due to experimental conditions was examined by using fura 2 in four different cell types responding to appropriate agonists when the cells were incubated in either CO2/HCO3 -- or HEPES-buffered media. Examined were: 1) the response of rat glomerular mesangial cells to arginine vasopressin, 2) the response of vascular smooth muscle cells to angiotensin II, 3) the response of adrenal glomerulosa cells to angiotensin II, and 4) the response of hypothalamic cells to insulin-like growth factor-1. In each cell type there was a significant difference in the pattern of agonist-induced change in [Ca2+](I) when HEPES vs. CO2/HCO3 - was used as the buffer system: in HEPES buffer, agonist addition led to a transient rise in [Ca2+](i) followed by a fall to a sustained plateau 27 to 34 nM higher than the original basal value, whereas in CO2/HCO3 - buffer, agonist addition led to an identical transient increase in [Ca2+](i) followed by a fall to a value within 10 nM or less of the preagonist level. The plateau value of [Ca2+](i) in the different buffers was examined in relationship to known differences in intracellular pH (pH(i)). It was found that measurements of [Ca2+](i) with fura 2 were influenced by shifts in pH(i) that occur when cells are incubated in either HEPES-buffered or CO2/HCO3 - media of differing pH0 values. However, at any given value of pH(i), the apparent [Ca2+](i) measured in cells incubated in HEPES-buffered media was slightly higher than in cells incubated in CO2/HCO3 - buffered media.

Original languageEnglish (US)
Pages (from-to)1638-1644
Number of pages7
JournalFASEB Journal
Volume4
Issue number6
StatePublished - Apr 25 1990
Externally publishedYes

Fingerprint

HEPES
Fura-2
Buffers
Angiotensin II
Cells
Arginine Vasopressin
Somatomedins
Fluorescent Dyes
Muscle
Rats
Zona Glomerulosa
Calcium
Mesangial Cells
Vascular Smooth Muscle
Smooth Muscle Myocytes
Cultured Cells

Keywords

  • HEPES
  • bicarbonate
  • calcium
  • cultured cells
  • fura 2

ASJC Scopus subject areas

  • Biotechnology
  • Biochemistry
  • Molecular Biology
  • Genetics

Cite this

Ganz, M. B., Rasmussen, J., Bollag, W. B., & Rasmussen, H. (1990). Effect of buffer systems and pH(i) on the measurement of [Ca2+](i) with fura 2. FASEB Journal, 4(6), 1638-1644.

Effect of buffer systems and pH(i) on the measurement of [Ca2+](i) with fura 2. / Ganz, M. B.; Rasmussen, J.; Bollag, Wendy B; Rasmussen, H.

In: FASEB Journal, Vol. 4, No. 6, 25.04.1990, p. 1638-1644.

Research output: Contribution to journalArticle

Ganz, MB, Rasmussen, J, Bollag, WB & Rasmussen, H 1990, 'Effect of buffer systems and pH(i) on the measurement of [Ca2+](i) with fura 2', FASEB Journal, vol. 4, no. 6, pp. 1638-1644.
Ganz, M. B. ; Rasmussen, J. ; Bollag, Wendy B ; Rasmussen, H. / Effect of buffer systems and pH(i) on the measurement of [Ca2+](i) with fura 2. In: FASEB Journal. 1990 ; Vol. 4, No. 6. pp. 1638-1644.
@article{6314181570944d00bdc790ca3973656d,
title = "Effect of buffer systems and pH(i) on the measurement of [Ca2+](i) with fura 2",
abstract = "The fluorescent probe, fura 2, is widely used to measure agonist-induced changes in intracellular calcium concentration ([Ca2+](i)) in cultured cells. However, in many instances, the results obtained in the same cell type have differed from one study to the next. The possibility that such differences might be due to experimental conditions was examined by using fura 2 in four different cell types responding to appropriate agonists when the cells were incubated in either CO2/HCO3 -- or HEPES-buffered media. Examined were: 1) the response of rat glomerular mesangial cells to arginine vasopressin, 2) the response of vascular smooth muscle cells to angiotensin II, 3) the response of adrenal glomerulosa cells to angiotensin II, and 4) the response of hypothalamic cells to insulin-like growth factor-1. In each cell type there was a significant difference in the pattern of agonist-induced change in [Ca2+](I) when HEPES vs. CO2/HCO3 - was used as the buffer system: in HEPES buffer, agonist addition led to a transient rise in [Ca2+](i) followed by a fall to a sustained plateau 27 to 34 nM higher than the original basal value, whereas in CO2/HCO3 - buffer, agonist addition led to an identical transient increase in [Ca2+](i) followed by a fall to a value within 10 nM or less of the preagonist level. The plateau value of [Ca2+](i) in the different buffers was examined in relationship to known differences in intracellular pH (pH(i)). It was found that measurements of [Ca2+](i) with fura 2 were influenced by shifts in pH(i) that occur when cells are incubated in either HEPES-buffered or CO2/HCO3 - media of differing pH0 values. However, at any given value of pH(i), the apparent [Ca2+](i) measured in cells incubated in HEPES-buffered media was slightly higher than in cells incubated in CO2/HCO3 - buffered media.",
keywords = "HEPES, bicarbonate, calcium, cultured cells, fura 2",
author = "Ganz, {M. B.} and J. Rasmussen and Bollag, {Wendy B} and H. Rasmussen",
year = "1990",
month = "4",
day = "25",
language = "English (US)",
volume = "4",
pages = "1638--1644",
journal = "FASEB Journal",
issn = "0892-6638",
publisher = "FASEB",
number = "6",

}

TY - JOUR

T1 - Effect of buffer systems and pH(i) on the measurement of [Ca2+](i) with fura 2

AU - Ganz, M. B.

AU - Rasmussen, J.

AU - Bollag, Wendy B

AU - Rasmussen, H.

PY - 1990/4/25

Y1 - 1990/4/25

N2 - The fluorescent probe, fura 2, is widely used to measure agonist-induced changes in intracellular calcium concentration ([Ca2+](i)) in cultured cells. However, in many instances, the results obtained in the same cell type have differed from one study to the next. The possibility that such differences might be due to experimental conditions was examined by using fura 2 in four different cell types responding to appropriate agonists when the cells were incubated in either CO2/HCO3 -- or HEPES-buffered media. Examined were: 1) the response of rat glomerular mesangial cells to arginine vasopressin, 2) the response of vascular smooth muscle cells to angiotensin II, 3) the response of adrenal glomerulosa cells to angiotensin II, and 4) the response of hypothalamic cells to insulin-like growth factor-1. In each cell type there was a significant difference in the pattern of agonist-induced change in [Ca2+](I) when HEPES vs. CO2/HCO3 - was used as the buffer system: in HEPES buffer, agonist addition led to a transient rise in [Ca2+](i) followed by a fall to a sustained plateau 27 to 34 nM higher than the original basal value, whereas in CO2/HCO3 - buffer, agonist addition led to an identical transient increase in [Ca2+](i) followed by a fall to a value within 10 nM or less of the preagonist level. The plateau value of [Ca2+](i) in the different buffers was examined in relationship to known differences in intracellular pH (pH(i)). It was found that measurements of [Ca2+](i) with fura 2 were influenced by shifts in pH(i) that occur when cells are incubated in either HEPES-buffered or CO2/HCO3 - media of differing pH0 values. However, at any given value of pH(i), the apparent [Ca2+](i) measured in cells incubated in HEPES-buffered media was slightly higher than in cells incubated in CO2/HCO3 - buffered media.

AB - The fluorescent probe, fura 2, is widely used to measure agonist-induced changes in intracellular calcium concentration ([Ca2+](i)) in cultured cells. However, in many instances, the results obtained in the same cell type have differed from one study to the next. The possibility that such differences might be due to experimental conditions was examined by using fura 2 in four different cell types responding to appropriate agonists when the cells were incubated in either CO2/HCO3 -- or HEPES-buffered media. Examined were: 1) the response of rat glomerular mesangial cells to arginine vasopressin, 2) the response of vascular smooth muscle cells to angiotensin II, 3) the response of adrenal glomerulosa cells to angiotensin II, and 4) the response of hypothalamic cells to insulin-like growth factor-1. In each cell type there was a significant difference in the pattern of agonist-induced change in [Ca2+](I) when HEPES vs. CO2/HCO3 - was used as the buffer system: in HEPES buffer, agonist addition led to a transient rise in [Ca2+](i) followed by a fall to a sustained plateau 27 to 34 nM higher than the original basal value, whereas in CO2/HCO3 - buffer, agonist addition led to an identical transient increase in [Ca2+](i) followed by a fall to a value within 10 nM or less of the preagonist level. The plateau value of [Ca2+](i) in the different buffers was examined in relationship to known differences in intracellular pH (pH(i)). It was found that measurements of [Ca2+](i) with fura 2 were influenced by shifts in pH(i) that occur when cells are incubated in either HEPES-buffered or CO2/HCO3 - media of differing pH0 values. However, at any given value of pH(i), the apparent [Ca2+](i) measured in cells incubated in HEPES-buffered media was slightly higher than in cells incubated in CO2/HCO3 - buffered media.

KW - HEPES

KW - bicarbonate

KW - calcium

KW - cultured cells

KW - fura 2

UR - http://www.scopus.com/inward/record.url?scp=0025357210&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=0025357210&partnerID=8YFLogxK

M3 - Article

C2 - 2318379

AN - SCOPUS:0025357210

VL - 4

SP - 1638

EP - 1644

JO - FASEB Journal

JF - FASEB Journal

SN - 0892-6638

IS - 6

ER -