Effects of Ca2+ agonists on cytosolic Ca2+ in isolated hepatocytes and on bile secretion in the isolated perfused rat liver.
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 mumol/L), the Ca2+ agonist vasopressin (10 nmol/L) and the Ca(2+)-mobilizing agent, 2,5-di(tert-butyl)-1,4-benzohydroquinone (25 mumol/L) on cytosolic Ca2+ in isolated hepatocytes 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 Ca(2+)-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 mumol/L) and the protein kinase C inhibitor H-7 (50 mumol/L) abolished the pressure increase but not the decrease in bile flow, whereas pretreatment with Ni2+ (25 mumol/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 ionophore-induced increase in Ca2+ was abolished completely by pretreatment of the hepatocytes with Ni2+, whereas the vasopressin-induced increase was reduced by 38%.(ABSTRACT TRUNCATED AT 400 WORDS)
Nathanson
M H
MH
Department of Internal Medicine, Yale University School of Medicine, New Haven, CT 06510.
Gautam
A
A
Bruck
R
R
Isales
C M
CM
Boyer
J L
JL
D000331:Aequorin; D000818:Animals; D001646:Bile / Q000378:metabolism*; D000001:Calcimycin / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism / Q000502:physiology*; D002469:Cell Separation; D003600:Cytosol / Q000378:metabolism*; D006873:Hydroquinones / Q000494:pharmacology; D066298:In Vitro Techniques; D008099:Liver / Q000166:cytology / Q000378:metabolism*; D008297:Male; D011312:Pressure; D051381:Rats; D011919:Rats, Inbred Strains; D014667:Vasopressins / Q000494:pharmacology
D006873:Hydroquinones; D014667:Vasopressins; C019359:2,5-di-tert-butylhydroquinone; D000001:Calcimycin; D000331:Aequorin; D002118:Calcium