Nuclear calcium gradients in cultured rat hepatocytes

M. M. Waybill, R. V. Yelamarty, Y. Zhang, R. C. Scaduto, K. F. LaNoue, C. J. Hsu, B. C. Smith, D. L. Tillotson, F. T.S. Yu, J. Y. Cheung

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Abstract

Ca2+ concentrations ([Ca2+]) in cytosol and nucleus in fura-2-loaded cultured rat hepatocytes were determined by three-dimensional (3-D) optical-sectioning microscopy. After determining the empirical 3-D point spread function of the fluorescence microscope-coupled digital video imaging system, contaminating light arising from optical planes above and below the plane of interest was removed by deconvolution using the nearest-neighboring approach (NNA) algorithm. Although deconvolution resulted in substantial improvement in accuracy of fluorescence intensity determinations in single-wavelength excitation images as well as sharper delineation of boundaries between cellular compartments, the complicated mathematical process did not significantly enhance the precision of [Ca2+] values derived from ratiometric (ratio of dual-wavelength excitation) images. In resting hepatocytes, cytosolic Ca2+ (210 ± 15 nM) was 1.6- to 2-fold higher than nuclear Ca2+ (128 ± 12 nM). This difference in Ca2+ between the two compartments was detected both in raw ratiometric images and in those processed with NNA algorithm. Addition of arginine vasopressin or epidermal growth factor resulted in significant increases (2- to 3-fold) in both cytosolic and nuclear Ca2+; however, the nuclear-to-cytosolic Ca2+ gradient was preserved in hepatocytes stimulated with mitogens. We conclude that the hepatocyte nuclear membrane contains Ca2+ permeability barriers and Ca2+ transport mechanisms that may be hormonally sensitive. We postulate that the increase in nuclear Ca2+ may be important in regulation of cell proliferation induced by mitogens, possibly by activating Ca2+-dependent endonucleases, nuclear calmodulin, or nuclear protein kinase C.

Original languageEnglish (US)
Pages (from-to)E49-E57
JournalAmerican Journal of Physiology - Endocrinology and Metabolism
Volume261
Issue number1 24-1
StatePublished - Jan 1 1991

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Hepatocytes
Calcium
Mitogens
Fluorescence
Arginine Vasopressin
Fura-2
Endonucleases
Nuclear Envelope
Calmodulin
Nuclear Proteins
Epidermal Growth Factor
Cytosol
Protein Kinase C
Microscopy
Permeability
Cell Proliferation
Light

All Science Journal Classification (ASJC) codes

  • Endocrinology, Diabetes and Metabolism
  • Physiology
  • Physiology (medical)

Cite this

Waybill, M. M., Yelamarty, R. V., Zhang, Y., Scaduto, R. C., LaNoue, K. F., Hsu, C. J., ... Cheung, J. Y. (1991). Nuclear calcium gradients in cultured rat hepatocytes. American Journal of Physiology - Endocrinology and Metabolism, 261(1 24-1), E49-E57.
Waybill, M. M. ; Yelamarty, R. V. ; Zhang, Y. ; Scaduto, R. C. ; LaNoue, K. F. ; Hsu, C. J. ; Smith, B. C. ; Tillotson, D. L. ; Yu, F. T.S. ; Cheung, J. Y. / Nuclear calcium gradients in cultured rat hepatocytes. In: American Journal of Physiology - Endocrinology and Metabolism. 1991 ; Vol. 261, No. 1 24-1. pp. E49-E57.
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Waybill, MM, Yelamarty, RV, Zhang, Y, Scaduto, RC, LaNoue, KF, Hsu, CJ, Smith, BC, Tillotson, DL, Yu, FTS & Cheung, JY 1991, 'Nuclear calcium gradients in cultured rat hepatocytes', American Journal of Physiology - Endocrinology and Metabolism, vol. 261, no. 1 24-1, pp. E49-E57.

Nuclear calcium gradients in cultured rat hepatocytes. / Waybill, M. M.; Yelamarty, R. V.; Zhang, Y.; Scaduto, R. C.; LaNoue, K. F.; Hsu, C. J.; Smith, B. C.; Tillotson, D. L.; Yu, F. T.S.; Cheung, J. Y.

In: American Journal of Physiology - Endocrinology and Metabolism, Vol. 261, No. 1 24-1, 01.01.1991, p. E49-E57.

Research output: Contribution to journalArticle

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T1 - Nuclear calcium gradients in cultured rat hepatocytes

AU - Waybill, M. M.

AU - Yelamarty, R. V.

AU - Zhang, Y.

AU - Scaduto, R. C.

AU - LaNoue, K. F.

AU - Hsu, C. J.

AU - Smith, B. C.

AU - Tillotson, D. L.

AU - Yu, F. T.S.

AU - Cheung, J. Y.

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N2 - Ca2+ concentrations ([Ca2+]) in cytosol and nucleus in fura-2-loaded cultured rat hepatocytes were determined by three-dimensional (3-D) optical-sectioning microscopy. After determining the empirical 3-D point spread function of the fluorescence microscope-coupled digital video imaging system, contaminating light arising from optical planes above and below the plane of interest was removed by deconvolution using the nearest-neighboring approach (NNA) algorithm. Although deconvolution resulted in substantial improvement in accuracy of fluorescence intensity determinations in single-wavelength excitation images as well as sharper delineation of boundaries between cellular compartments, the complicated mathematical process did not significantly enhance the precision of [Ca2+] values derived from ratiometric (ratio of dual-wavelength excitation) images. In resting hepatocytes, cytosolic Ca2+ (210 ± 15 nM) was 1.6- to 2-fold higher than nuclear Ca2+ (128 ± 12 nM). This difference in Ca2+ between the two compartments was detected both in raw ratiometric images and in those processed with NNA algorithm. Addition of arginine vasopressin or epidermal growth factor resulted in significant increases (2- to 3-fold) in both cytosolic and nuclear Ca2+; however, the nuclear-to-cytosolic Ca2+ gradient was preserved in hepatocytes stimulated with mitogens. We conclude that the hepatocyte nuclear membrane contains Ca2+ permeability barriers and Ca2+ transport mechanisms that may be hormonally sensitive. We postulate that the increase in nuclear Ca2+ may be important in regulation of cell proliferation induced by mitogens, possibly by activating Ca2+-dependent endonucleases, nuclear calmodulin, or nuclear protein kinase C.

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Waybill MM, Yelamarty RV, Zhang Y, Scaduto RC, LaNoue KF, Hsu CJ et al. Nuclear calcium gradients in cultured rat hepatocytes. American Journal of Physiology - Endocrinology and Metabolism. 1991 Jan 1;261(1 24-1):E49-E57.