Possible role of Pi supply in mitochondrial actions of glucagon

Abstract

Glucagon is able to diminish the net release of inorganic phosphate (Pi) occurring on incubation of isolated hepatocytes from 48-h-starved rats. Concomitantly the hormone increases the cellular Pi content. This is associated with a rise of Pi in the cytosolic fraction. Other hormonal effectors like phenylephrine, vasopressin and angiotensin I1 exert a smaller and transient effect as compared to glucagon. It is proposed that this increase in Pi availability to the mitochondria, by favouring substratc lcvcl phosphorylation at the succinyl-CoA synthetasc step plays a role in the development of the metabolite pattern found in the mitochondrial matrix spacc after exposure of hepatocytes to glucagon or the above agents. With regard to the glutamate level this view is evidcnced by the finding that its hormone-dependent decrease was inversely correlated to the respective increase in the cytosolic Pi concentration. Further evidence is provided by experiments with isolated mitochondria incubatcd under state-3 conditions at medium Pi concentrations corresponding to those metabolically active in the cytosolic compartment of control and glucagon-stiinulatcd hepatocytes, being 2 mM and 3 mM, respectively. Increasing medium phosphate concentration from 2 mM to 3 mM caused a marked decrease in the levcl of succinyl-CoA and increased the rates of 2-oxoglutarate utilization and of malate and phosphoenolpyruvate production. Citrulline synthesis also was found to be stimulated at 3 tnM Pi. Taken together our results suggest a rolc of Pi supply in mitochondrial actions of glucagon in intact hepatocytes. Moreover, they could contribute to a bcttcr interpretation of glucagon effects on isolated mitochondria from hormone-pretreated liver cells.The effects of glucagon on liver mitochondria may be divided into two groups: the first one comprising the changes occurring in the mitochondrial compartment of intact hepatocytes [l] as revealed by the digitonin fractionation technique [2] and the second one consisting of the effects demonstrable after isolation of the mitochondria from glucagon-stimulated hepatocytes. With respect to the first group the drop in glutamate and 2-oxoglutarate as well as thc increase in the ATP, phosphoenolpyruvate and acetyl-CoA contents [l] are noteworthy. As to the second group about 15 different parameters have been described out of which the apparent stimulation of respiration (for review scc [3]) and the activation of pyruvate carboxylase and citrullinc formation [9,181 have been extensively investigated. The results have been interpreted as to indicate that the isolated mitochondria represent a valid model for the hormone's action on mitochondrial metabolism in intact hepatocytes. Howcver, evidence has been presented for the fact that a number of the hormone effects demonsti-ablc with the isolated mitochondria strongly depend on the conditions of mitochondria isolation, storage and incubation [I 91. In the course of studies on the influence of various isolation media on the preservation of functional in...

Stimulation by 3-hydroxybutyrate of pyruvate carboxylation in mitochondria from rat liver

“…Regarding the latter the matrix concentration has been determined to amount to about 0.8 mM in liver cells from fasted rats incubated with lactate as the substrate [Is, 351, being increased up to 3 mM by the addition of oleate (0.4 mM) [35]. This finding and the 3-hydroxybutyrate dependence of pyruvate carboxylation (Fig.…”

“…With respect to the question as to whether the 3-hydroxybutyrate effect might have regulatory relevance for pyruvate carboxylation in intact liver cells it is noteworthy that effective changes in the 3-hydroxybutyrate/acetoacetate ratio and in the concentration of 3-hydroxybutyrate occur in the mitochondrial compartment [35]. Regarding the latter the matrix concentration has been determined to amount to about 0.8 mM in liver cells from fasted rats incubated with lactate as the substrate [Is, 351, being increased up to 3 mM by the addition of oleate (0.4 mM) [35].…”

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

“…The activation of hepatic oxoglutarate dehydrogenase by vasopressin has been demonstrated to be dependent on Ca2 ' [20]. Alternatively, the hormonal activation of oxoglutarate dehydrogenase may be due to a decrease in succinyl-CoA concentration as a consequence of an increase in mitochondrial phosphate concentration [21]. Exactly how a hormone employing cyclic AMP as a second messenger causes an activation of the Ca2+-sensitive dehydrogenases remains to be established.…”

“…Activation of phosphorylase by glucagon is mediated by an increase in cyclic AMP concentration. Ionic signals arc also involved in the regulation of liver phosphorylase activity: vasopressin, for example, acts independently of an increase in cyclic AMP concentration [I] succinyl-CoA concentration as a consequence of an increase in mitochondrial phosphate concentration [21]. Exactly how a hormone employing cyclic AMP as a second messenger causes an activation of the Ca2+-sensitive dehydrogenases remains to be established.…”

Phosphate and calcium uptake by mitochondria and by perfused rat liver induced by the synergistic action of glucagon and vasopressin

“…On the other hand, there is evidence to suggest that glucagon, cyclic AMP and adrenaline are able to stimulate Pi exchange in perfused rat liver (De Venanzi et al, 1974) and in perfused rat heart (Medina & Illingworth, 1984) and that glucagon administration to hepatocytes induces a quite rapid increase in cellular Pi content (Siess et al, 1984). Moreover, administration of glucagon to the whole animal or to perfused rat liver induces an increase in the Pi content of the subsequently isolated mitochondria (Prpic et al, 1978;Hughes & Barritt, 1978).…”