Hypoxia inhibits cyclic nucleotide-stimulated epithelial ion transport: Role for nucleotide cyclases as oxygen sensors

Cormac T. Taylor, Steven J. Lisco, Christopher S. Awtrey, Sean P. Colgan

Research output: Contribution to journalArticle

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Abstract

Decreased oxygen delivery to cells (hypoxia) is prevalent in a number of important diseases. Little is known about mechanisms of oxygen sensing at the cellular level or about whether functional correlates of oxygen sensing exist. In this study, we examined the impact of hypoxia on stimulated epithelial ion transport function. T84 cells, a model of intestinal epithelia, were grown on permeable supports, exposed to hypoxia (range 1-21% O2) for periods of time between 0 and 72 h and assessed for stimulated ion transport. Hypoxia evoked a specific decrease in cyclic nucleotide-stimulated (cAMP and cGMP) but not Ca++-stimulated ion transport. 86Rb (K+ tracer) uptake and 125I (Cl- tracer) efflux were reduced in hypoxic cells by >50% and >40%, respectively, fluid movement was reduced by hypoxia (>50% decrease) and reoxygenation resulted in partial recovery of the ion transport responses. Stimulated and basal levels of both cAMP and cGMP were decreased in response to hypoxia, although intracellular ATP levels were unaltered under similar conditions. Exogenous addition of cobalt, nickel or manganese, all of which compete for oxygen binding on hemecontaining proteins, mimicked hypoxia. Because guanylate cyclase is a heme protein, we measured the influence of cobalt on activity of guanylate cyclase in purified plasma membrane preparations and found cobalt to inhibit stimulated cGMP levels in this cell-free system. Finally, pharmacological lowering of intracellular cGMP (using LY83583) resulted in decreased cAMP-stimulated Cl- secretion, and direct elevation of cGMP (using 8-bromo-cGMP or dibutyryl-cGMP) restored this hypoxia-induced activity. We conclude that a potential oxygen-sensing mechanism of epithelial cells involves the cooperation of heme-containing proteins such as guanylate cyclase and that biochemical cross-talk between cAMP- and cGMP-stimulated pathways may be important in such responses.

Original languageEnglish (US)
Pages (from-to)568-575
Number of pages8
JournalJournal of Pharmacology and Experimental Therapeutics
Volume284
Issue number2
StatePublished - Feb 1 1998

Fingerprint

Cyclic Nucleotides
Ion Transport
Nucleotides
Oxygen
Guanylate Cyclase
Cobalt
6-anilino-5,8-quinolinedione
Hemeproteins
Cell Hypoxia
Cell-Free System
Intestinal Mucosa
Manganese
Hypoxia
Nickel
Heme
Proteins
Adenosine Triphosphate
Epithelial Cells
Cell Membrane
Pharmacology

ASJC Scopus subject areas

  • Molecular Medicine
  • Pharmacology

Cite this

Hypoxia inhibits cyclic nucleotide-stimulated epithelial ion transport : Role for nucleotide cyclases as oxygen sensors. / Taylor, Cormac T.; Lisco, Steven J.; Awtrey, Christopher S.; Colgan, Sean P.

In: Journal of Pharmacology and Experimental Therapeutics, Vol. 284, No. 2, 01.02.1998, p. 568-575.

Research output: Contribution to journalArticle

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abstract = "Decreased oxygen delivery to cells (hypoxia) is prevalent in a number of important diseases. Little is known about mechanisms of oxygen sensing at the cellular level or about whether functional correlates of oxygen sensing exist. In this study, we examined the impact of hypoxia on stimulated epithelial ion transport function. T84 cells, a model of intestinal epithelia, were grown on permeable supports, exposed to hypoxia (range 1-21{\%} O2) for periods of time between 0 and 72 h and assessed for stimulated ion transport. Hypoxia evoked a specific decrease in cyclic nucleotide-stimulated (cAMP and cGMP) but not Ca++-stimulated ion transport. 86Rb (K+ tracer) uptake and 125I (Cl- tracer) efflux were reduced in hypoxic cells by >50{\%} and >40{\%}, respectively, fluid movement was reduced by hypoxia (>50{\%} decrease) and reoxygenation resulted in partial recovery of the ion transport responses. Stimulated and basal levels of both cAMP and cGMP were decreased in response to hypoxia, although intracellular ATP levels were unaltered under similar conditions. Exogenous addition of cobalt, nickel or manganese, all of which compete for oxygen binding on hemecontaining proteins, mimicked hypoxia. Because guanylate cyclase is a heme protein, we measured the influence of cobalt on activity of guanylate cyclase in purified plasma membrane preparations and found cobalt to inhibit stimulated cGMP levels in this cell-free system. Finally, pharmacological lowering of intracellular cGMP (using LY83583) resulted in decreased cAMP-stimulated Cl- secretion, and direct elevation of cGMP (using 8-bromo-cGMP or dibutyryl-cGMP) restored this hypoxia-induced activity. We conclude that a potential oxygen-sensing mechanism of epithelial cells involves the cooperation of heme-containing proteins such as guanylate cyclase and that biochemical cross-talk between cAMP- and cGMP-stimulated pathways may be important in such responses.",
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