Effect of Ca²⁺ agonists in the perfused liver: Determination via laser scanning confocal microscopy

Ca²⁺ is a critical intracellular second messenger, but few studies have examined Ca²⁺ signaling in whole organs. The amplitude and frequency of Ca²⁺ oscillations encode important cellular information. Using laser scanning confocal microscopy in the indo 1 acetoxymethyl ester dye-loaded rat liver, we investigated the effect of various Ca²⁺ agonists that act at distinct mechanistic sites on Ca²⁺ signaling. Perfusion with supra- threshold doses of arginine vasopressin (AVP) (2-20 nM) caused a single Ca²⁺ wave that originated in the pericentral vein region and spread centrifugally to the periportal area. Lower doses of AVP (0.2-2 nM) caused multiple Ca²⁺ waves and Ca²⁺ oscillations. Perfusion with ATP (1.4-17.5 μM) caused rapid transient elevations in intracellular free Ca²⁺ concentration ([Ca²⁺](i)) occurring in isolated hepatocytes or groups of hepatocytes throughout the lobule and were of shorter duration than those due to AVP. Also in contrast to AVP, there was no specific anatomic location within the hepatic lobule that was more susceptible to ATP. Thapsigargin and cyclopiazonic acid did not cause a Ca²⁺ wave but rather produced a uniform and fairly simultaneous increase in [Ca²⁺](i) in all hepatocytes in the lobule. Perfusion with 14 μM ryanodine produced a single transient spike in [Ca²⁺](i) in a small number (<2%) of hepatocytes. Dantrolene, an inhibitor of Ca²⁺ release, reduced the increased [Ca²⁺](i) occurring after AVP. Insight into the mechanism of action of these Ca²⁺-active compounds on Ca²⁺ signaling in the intact liver is provided.