Effects of ATP and adenosine addition on activity of oxoglutarate dehydrogenase and the concentration of cytoplasmic free Ca2+ in rat hepatocytes.

Abstract

Addition of ATP (100 microM) to hepatocytes from starved rats incubated with 5 mM [1-14C]glutamine caused a stimulation of glucose formation; the magnitude of the concomitant increases in 14CO2 production and glutamine consumption indicate that flux from glutamine to glucose was increased. ATP also caused a simultaneous decrease in the cell content of oxoglutarate; together with the increased flux this is consistent with an activation of oxoglutarate dehydrogenase. In corroboration of this, a stimulation by ATP of gluconeogenesis and a decrease in oxoglutarate was also observed with 5 mM proline as substrate. ATP caused an increase in hepatocyte cytoplasmic free Ca2+ concentration, [Ca2+]c, as indicated by the increase in the fluorescence of cytoplasmically trapped quin2, from a resting value of about 0.2 microM to greater than 1 microM. The mechanism of oxoglutarate dehydrogenase activation may be via an increase in mitochondrial Ca2+ content as a consequence of the increase in [Ca2+]c. The effects of 100 microM adenosine were also investigated. An increase in flux from glutamine to glucose was observed together with a decrease in the cell oxoglutarate, thus indicating that adenosine addition to hepatocytes could also activate oxoglutarate dehydrogenase. The activation by adenosine was less than that produced by ATP. Adenosine caused a small apparent increase in [Ca2+]c to 0.3-0.4 microM; it remains to be established if this effect, which is small relative to that of ATP, is sufficient to elicit the activation of oxoglutarate dehydrogenase: alternative mechanisms may exist.