Effects of Ca2+ agonists on cytosolic Ca2+ in isolated hepatocytes and on bile secretion in the isolated perfused rat liver

Michael H. Nathanson, Anil Gautam, Rafael Bruck, Carlos M Isales, James L. Boyer

Research output: Contribution to journalArticle

63 Citations (Scopus)

Abstract

The effects of increases in cytosolic Ca2+ on hepatocyte bile secretion are unknown. A number of agents that alter levels of cytosolic Ca2+ in the hepatocyte also produce hepatic vasoconstriction and activate protein kinase C, which complicates interpretations of their effects on bile secretion. To better understand the role of cytosolic Ca2+ in bile secretion, we examined the effect of the Ca2+ ionophore A23187 (0.1 μmol/L), the Ca2+ agonist vasopressin (10 nmol/L) and the Ca2+‐mobilizing agent, 2,5‐di(tert‐butyl)‐1,4‐benzohydroquinone (25 μmol/L) on cytosolic Ca2+ in isolated hepatooytes and on bile flow in the isolated perfused rat liver, using vasodilators and inhibitors of protein kinase C and Ca2+ influx. Single‐pass perfused livers were used, and cytosolic Ca2+ was measured by luminescent photometry in isolated hepatocytes loaded with the Ca2+‐sensitive photoprotein aequorin. After A23187 perfusion, a sustained 74% ± 10% (mean ± S. D.) decrease in bile flow and a sustained 271% ± 50% increase in perfusion pressure was observed. Simultaneous pretreatment with the vasodilator papaverine (25 μmol/L) and the protein kinase C inhibitor H‐7 (50 μmol/L) abolished the pressure in crease but not the decrease in bile flow, whereas pretreatment with Ni2+ (25 μmol/L) to block the influx of extracellular Ca2+ markedly reduced both the pressure increase and the decrease in bile flow. Vasopressin produced a transient (mean = 6 min) 75% ± 4% decrease in bile flow and a sustained 7% ± 4% increase in perfusion pressure. Pretreatment with H‐7 alone corrected the vasopressin‐induced pressure increase but also failed to eliminate the decrease in bile flow, whereas pretreatment with Ni2+ decreased the magnitude of the decrease by two‐thirds without affecting the increase in perfusion pressure. 2,5′‐di(tert‐butyl)‐1,4‐benzohydroquinone produced a transient 65% ± 20% decrease in bile flow and a transient 56% ± 15% increase in perfusion pressure. In isolated hepatocytes, bromo‐A23187, the nonfluorescent form of the ionophore, produced a sustained 56% ± 32% increase in the cytosolic Ca2+ signal, whereas vasopressin resulted in a transient 241% ± 75% increase and 2,5‐di(tert‐butyl)‐1,4‐benzohydroquinone resulted in a sustained 149% ± 66% increase. The ionophoreinduced increase in Ca2+ was abolished completely by pretreatment of the hepatocytes with Ni2+, whereas the vasopressin‐induced increase was reduced by 38%. These results indicate that agents that increase cytosolic Ca2+ in isolated hepatocytes from either internal or external sources also inhibit bile secretion in the isolated perfused liver independently of hemodynamic or protein kinase C effects. Furthermore, conditions in which the Ca2+ rise is inhibited in isolated hepatocytes lead to decreased inhibition of bile secretion in the perfused liver. These observations, along with the temporal relationship between changes in cytosolic Ca2+ in isolated hepatocytes and decreased bile flow in isolated perfused rat livers, suggest that increased cytosolic Ca2+ may play an inhibitory role in the regulation of bile secretion. (HEPATOLOGY 1992;15:107‐116).

Original languageEnglish (US)
Pages (from-to)107-116
Number of pages10
JournalHepatology
Volume15
Issue number1
DOIs
StatePublished - Jan 1 1992

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Bile
Hepatocytes
Liver
Pressure
Perfusion
Protein Kinase C
Vasopressins
Ionophores
Calcimycin
Vasodilator Agents
Luminescent Proteins
Aequorin
Photometry
Papaverine
Protein C Inhibitor
Protein Kinase Inhibitors
Vasoconstriction
Hemodynamics

ASJC Scopus subject areas

  • Hepatology

Cite this

Effects of Ca2+ agonists on cytosolic Ca2+ in isolated hepatocytes and on bile secretion in the isolated perfused rat liver. / Nathanson, Michael H.; Gautam, Anil; Bruck, Rafael; Isales, Carlos M; Boyer, James L.

In: Hepatology, Vol. 15, No. 1, 01.01.1992, p. 107-116.

Research output: Contribution to journalArticle

Nathanson, Michael H. ; Gautam, Anil ; Bruck, Rafael ; Isales, Carlos M ; Boyer, James L. / Effects of Ca2+ agonists on cytosolic Ca2+ in isolated hepatocytes and on bile secretion in the isolated perfused rat liver. In: Hepatology. 1992 ; Vol. 15, No. 1. pp. 107-116.
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T1 - Effects of Ca2+ agonists on cytosolic Ca2+ in isolated hepatocytes and on bile secretion in the isolated perfused rat liver

AU - Nathanson, Michael H.

AU - Gautam, Anil

AU - Bruck, Rafael

AU - Isales, Carlos M

AU - Boyer, James L.

PY - 1992/1/1

Y1 - 1992/1/1

N2 - The effects of increases in cytosolic Ca2+ on hepatocyte bile secretion are unknown. A number of agents that alter levels of cytosolic Ca2+ in the hepatocyte also produce hepatic vasoconstriction and activate protein kinase C, which complicates interpretations of their effects on bile secretion. To better understand the role of cytosolic Ca2+ in bile secretion, we examined the effect of the Ca2+ ionophore A23187 (0.1 μmol/L), the Ca2+ agonist vasopressin (10 nmol/L) and the Ca2+‐mobilizing agent, 2,5‐di(tert‐butyl)‐1,4‐benzohydroquinone (25 μmol/L) on cytosolic Ca2+ in isolated hepatooytes and on bile flow in the isolated perfused rat liver, using vasodilators and inhibitors of protein kinase C and Ca2+ influx. Single‐pass perfused livers were used, and cytosolic Ca2+ was measured by luminescent photometry in isolated hepatocytes loaded with the Ca2+‐sensitive photoprotein aequorin. After A23187 perfusion, a sustained 74% ± 10% (mean ± S. D.) decrease in bile flow and a sustained 271% ± 50% increase in perfusion pressure was observed. Simultaneous pretreatment with the vasodilator papaverine (25 μmol/L) and the protein kinase C inhibitor H‐7 (50 μmol/L) abolished the pressure in crease but not the decrease in bile flow, whereas pretreatment with Ni2+ (25 μmol/L) to block the influx of extracellular Ca2+ markedly reduced both the pressure increase and the decrease in bile flow. Vasopressin produced a transient (mean = 6 min) 75% ± 4% decrease in bile flow and a sustained 7% ± 4% increase in perfusion pressure. Pretreatment with H‐7 alone corrected the vasopressin‐induced pressure increase but also failed to eliminate the decrease in bile flow, whereas pretreatment with Ni2+ decreased the magnitude of the decrease by two‐thirds without affecting the increase in perfusion pressure. 2,5′‐di(tert‐butyl)‐1,4‐benzohydroquinone produced a transient 65% ± 20% decrease in bile flow and a transient 56% ± 15% increase in perfusion pressure. In isolated hepatocytes, bromo‐A23187, the nonfluorescent form of the ionophore, produced a sustained 56% ± 32% increase in the cytosolic Ca2+ signal, whereas vasopressin resulted in a transient 241% ± 75% increase and 2,5‐di(tert‐butyl)‐1,4‐benzohydroquinone resulted in a sustained 149% ± 66% increase. The ionophoreinduced increase in Ca2+ was abolished completely by pretreatment of the hepatocytes with Ni2+, whereas the vasopressin‐induced increase was reduced by 38%. These results indicate that agents that increase cytosolic Ca2+ in isolated hepatocytes from either internal or external sources also inhibit bile secretion in the isolated perfused liver independently of hemodynamic or protein kinase C effects. Furthermore, conditions in which the Ca2+ rise is inhibited in isolated hepatocytes lead to decreased inhibition of bile secretion in the perfused liver. These observations, along with the temporal relationship between changes in cytosolic Ca2+ in isolated hepatocytes and decreased bile flow in isolated perfused rat livers, suggest that increased cytosolic Ca2+ may play an inhibitory role in the regulation of bile secretion. (HEPATOLOGY 1992;15:107‐116).

AB - The effects of increases in cytosolic Ca2+ on hepatocyte bile secretion are unknown. A number of agents that alter levels of cytosolic Ca2+ in the hepatocyte also produce hepatic vasoconstriction and activate protein kinase C, which complicates interpretations of their effects on bile secretion. To better understand the role of cytosolic Ca2+ in bile secretion, we examined the effect of the Ca2+ ionophore A23187 (0.1 μmol/L), the Ca2+ agonist vasopressin (10 nmol/L) and the Ca2+‐mobilizing agent, 2,5‐di(tert‐butyl)‐1,4‐benzohydroquinone (25 μmol/L) on cytosolic Ca2+ in isolated hepatooytes and on bile flow in the isolated perfused rat liver, using vasodilators and inhibitors of protein kinase C and Ca2+ influx. Single‐pass perfused livers were used, and cytosolic Ca2+ was measured by luminescent photometry in isolated hepatocytes loaded with the Ca2+‐sensitive photoprotein aequorin. After A23187 perfusion, a sustained 74% ± 10% (mean ± S. D.) decrease in bile flow and a sustained 271% ± 50% increase in perfusion pressure was observed. Simultaneous pretreatment with the vasodilator papaverine (25 μmol/L) and the protein kinase C inhibitor H‐7 (50 μmol/L) abolished the pressure in crease but not the decrease in bile flow, whereas pretreatment with Ni2+ (25 μmol/L) to block the influx of extracellular Ca2+ markedly reduced both the pressure increase and the decrease in bile flow. Vasopressin produced a transient (mean = 6 min) 75% ± 4% decrease in bile flow and a sustained 7% ± 4% increase in perfusion pressure. Pretreatment with H‐7 alone corrected the vasopressin‐induced pressure increase but also failed to eliminate the decrease in bile flow, whereas pretreatment with Ni2+ decreased the magnitude of the decrease by two‐thirds without affecting the increase in perfusion pressure. 2,5′‐di(tert‐butyl)‐1,4‐benzohydroquinone produced a transient 65% ± 20% decrease in bile flow and a transient 56% ± 15% increase in perfusion pressure. In isolated hepatocytes, bromo‐A23187, the nonfluorescent form of the ionophore, produced a sustained 56% ± 32% increase in the cytosolic Ca2+ signal, whereas vasopressin resulted in a transient 241% ± 75% increase and 2,5‐di(tert‐butyl)‐1,4‐benzohydroquinone resulted in a sustained 149% ± 66% increase. The ionophoreinduced increase in Ca2+ was abolished completely by pretreatment of the hepatocytes with Ni2+, whereas the vasopressin‐induced increase was reduced by 38%. These results indicate that agents that increase cytosolic Ca2+ in isolated hepatocytes from either internal or external sources also inhibit bile secretion in the isolated perfused liver independently of hemodynamic or protein kinase C effects. Furthermore, conditions in which the Ca2+ rise is inhibited in isolated hepatocytes lead to decreased inhibition of bile secretion in the perfused liver. These observations, along with the temporal relationship between changes in cytosolic Ca2+ in isolated hepatocytes and decreased bile flow in isolated perfused rat livers, suggest that increased cytosolic Ca2+ may play an inhibitory role in the regulation of bile secretion. (HEPATOLOGY 1992;15:107‐116).

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