Abstract

Extracellular ATP, ADP and GTP increased the intracellular free Ca2+ concentration ([Ca2+]i) in a suspension of isolated rat hepatocytes. The [Ca2+]i was determined by measuring fura-2 fluorescence, and its increase was biphasic. The initial transient rise was followed by a longer-lasting plateau. The peak of the early component preceded the plateau level of the second component. A time course of change in [Ca2+]i in single cells at 100 microM-ATP was very similar to that observed in the suspension system. Preincubation of hepatocytes with 40 mM-caffeine, 2 mM-oxalate or 60 microM-dantrolene sodium inhibited the P2 purinergic response. The plateau phase was not observed when measured in the presence of extracellular 100 microM-LaCl3 or in the absence of extracellular Ca2+. The distribution of [Ca2+]i in single hepatocytes was also determined by fluorescence image analysis. In the initial phase, the increase in [Ca2+]i is greater in the peripheral region than the central region of the cell. Degradation of extracellular ATP by ecto-ATPase in the hepatocyte suspension was measured; the amount of ATP degradation was less than 10-15% of the initial amount (100 microM) during the measurement of the intracellular [Ca2+]i in the cell suspension. Extracellular ATP stimulated glucose synthesis. The rate of glucose production also showed two components, the initial fast component within 1 min and the subsequent slower component. The rate of the initial fast component did not depend on the presence or absence of extracellular Ca2+, whereas the rate of the subsequent component depended on it. The present study shows that the initial transient rise in [Ca2+]i plays an important role in triggering the gluconeogenesis.

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