Intracellular calcium in the isolated rat liver: correlation to glucose release, K(+) balance and bile flow.

This study correlates whole organ measurements of intracellular calcium concentration ([Ca(2+)](i)) with hormone-induced (epinephrine, vasopressin) changes of liver functions (glucose release, K(+) balance and bile flow). [Ca(2+)](i) was measured in the isolated perfused rat liver using the sensor Fura-2 and applying liver surface fluorescence spectroscopy. The technique was improved by (i) minimizing biliary elimination of the sensor by employing a rat strain deficient in canalicular organic anion transport (TR(-) mutation) and (ii) by correcting for changes of interfering intrinsic organ fluorescence that was shown to depend on the oxidation-reduction state (NAD(P)H content) of the organ. Epinephrine (50 nM) elicits an instantaneous peak rise of [Ca(2+)](i) to approx. 400 nM, followed by a sustained elevation that depends on the presence of extracellular Ca(2+). The rise of [Ca(2+)](i) coincides with initiation of glucose release, transient K(+) uptake, and transient stimulation of bile flow. Vasopressin (2 nM) exerts qualitatively similar effects. The transient rise of bile flow is attributed to Ca(2+)-mediated contraction of the pericanalicular actin-myosin web of hepatocytes.

Effects of insulin-like growth factor I on basal and stimulated glucose fluxes in rat liver.

Effects of insulin-like growth factor I (IGF-I) and insulin on glucose and potassium fluxes were examined by measuring transhepatic glucose and potassium balance in isolated perfused rat livers. At 1 nM, both IGF-I and insulin decreased basal glucose release by approximately 64% (P < 0.05). Adrenaline (epinephrine)-stimulated glucose release (42.6 +/- 4.5 micromol/g of liver within 30 min) was inhibited (P < 0.05) by approximately 32 and approximately 52% during IGF-I and insulin exposure, which was accompanied by reduced cAMP release (-71 and -80%, P < 0.05). IGF-I- and insulin-induced reduction of glucose release only decreased during calcium-free perfusion, but not during inhibition of phosphoinositide 3-kinase by wortmannin. Both IGF-I and insulin induced net potassium uptake, while insulin also attenuated the response to adrenaline. In conclusion, IGF-I causes (i) insulin-like inhibition of hepatic glycogenolysis, even at low, nanomolar concentrations, which is associated with decreased cAMP release, reduced in the absence of Ca(2+), but not mediated by phosphoinositide 3-kinase, (ii) reduction of adrenaline-induced glycogenolysis and (iii) net potassium uptake under basal conditions.

Acute troglitazone action in isolated perfused rat liver.

1. The thiazolidinedione compound, troglitazone, enhances insulin action and reduces plasma glucose concentrations when administered chronically to type 2 diabetic patients. 2. To analyse to what extent thiazolidinediones interfere with liver function, we examined the acute actions of troglitazone (0.61 and 3.15 microM) on hepatic glucose and lactate fluxes, bile secretion, and portal pressure under basal, insulin- and/or glucagon-stimulated conditions in isolated perfused rat livers. 3. During BSA-free perfusion, high dose troglitazone increased basal (P < 0.01), but inhibited glucagon-stimulated incremental glucose production by approximately 75% (10.0 +/- 2.5 vs control: 40.0 +/- 7.2 micromol g liver(-1), P < 0.01). In parallel, incremental lactate release rose approximately 6 fold (13.1 +/- 5.9 vs control: 2.2 +/- 0.8 mmol g liver(-1), P < 0.05), while bile secretion declined by approximately 67% [0.23 +/- 0.02 vs control: 0.70 +/- 0.05 mg g liver(-1) min(-1)), P < 0.001]. Low dose troglitazone infusion did not enhance the inhibitory effect of insulin on glucagon-stimulated glucose production, but rapidly increased lactate release (P < 0.0005) and portal venous pressure (+0.17 +/- 0.07 vs +0.54 +/- 0.07 cm buffer height, P < 0.0001). 4. These results indicate that troglitazone exerts both insulin-like and non-insulin-like hepatic effects, which are blunted by addition of albumin, possibly due to troglitazone binding.

Acute effect of leptin on hepatic glycogenolysis and gluconeogenesis in perfused rat liver.

Leptin circulates in blood and is involved in body weight control primarily via hypothalamic receptors. To examine its direct metabolic action, effects of short-term portal leptin infusion: 1) on postprandial basal and epinephrine-stimulated glycogenolysis; and 2) on postabsorptive lactate-stimulated gluconeogenesis were studied in isolated perfused rat livers. Incremental epinephrine (150 pmol x min-1 x g-1 liver)-stimulated glucose release (in micromol/g liver within 30 minutes; control: 28.3 +/- 2.8) was suppressed (P <.05) by 44% (15.8 +/- 1.6), by 48% (14.6 +/- 4.1), and by 53% (13.3 +/- 2.1) during insulin (3 pmol x min-1 x g-1 liver), leptin (30 pmol x min-1 x g-1 liver), and simultaneous leptin + insulin infusion. Perfusate cyclic adenosine monophosphate increased approximately twofold during epinephrine stimulation in all groups. Neither leptin nor insulin affected hepatic lactate production, bile flow, or portal pressure in the fed state. In the postabsorptive state (20-hour fasting), rates of lactate (10 mmol/L)-dependent hepatic glucose release (in micromol. min-1 x g-1 liver; control: 0.12 +/- 0.01) were increased (P <.01) to 0.35 +/- 0.02 and to 0.24 +/- 0.01 by glucagon (3 pmol x min-1 x g-1 liver) and by leptin (15 pmol x min-1 x g-1 liver), respectively. In parallel, lactate uptake rates (in micromol x min-1 x g-1 liver) were higher in the presence of both glucagon (0.90 +/- 0. 03) and leptin (0.84 +/- 0.02) compared with control (0.68 +/- 0.04). In conclusion, leptin modulates hepatic glucose fluxes and may contribute to direct humoral regulation of liver glycogen stores in the fasted as well as in the fed state.

Failure of leptin to affect basal and insulin-stimulated glucose metabolism of rat skeletal muscle in vitro.

Studies on different isolated tissues have provided evidence that leptin may directly modulate cellular glucose handling. The present study was performed to elucidate leptin's action on basal and insulin-stimulated glucose metabolism in native muscle tissue, which under physiological circumstances is the quantitatively most important target tissue of insulin. Isolated rat soleus muscle strips were incubated for 1 h in the absence or presence of leptin (0, 1, 10, or 100 nmol/l) under basal or insulin-stimulated conditions (10 nmol/l). No effects of leptin were found on the rates of 3H-2-deoxy-glucose transport (basal: control, 314+/-14; 1 nmol/l leptin, 320+/-17; 10 nmol/l leptin, 314+/-13; 100 nmol/l leptin, 322+/-16; insulin-stimulated: control, 690+/-33; 1 nmol/l leptin, 691+/-29; 10 nmol/l leptin, 665+/-26; 100 nmol/l leptin, 664+/-27; cpm x mg(-1) x h(-1); NS vs respective control) and on net glucose incorporation into glycogen (basal: control, 1.75+/-0.18; 1 nmol/l leptin, 2.01+/-0.13; 10 nmol/l leptin, 1.92+/-0.11; 100 nmol/l leptin, 1.81+/-0.13; insulin-stimulated: control, 5.98+/-0.40; 1 nmol/l leptin, 5.93+/-0.30; 10 nmol/l leptin, 5.46+/-0.25; 100 nmol/l leptin, 5.85+/-0.30; micromol x g(-1) x h(-1); NS vs respective control). In parallel, leptin failed to affect rates of aerobic and anaerobic glycolysis as well as muscle glycogen content. Further experiments revealed that the inability of leptin to directly affect muscle glucose handling prevailed independently of muscle fiber type (soleus and epitrochlearis muscle), of ambient insulin concentrations (0-30 nmol/l), and of leptin exposure time (1 h or 6 h). Thus, our findings fail to support speculations about a physiological role of direct insulin-mimetic or insulin-desensitizing effects of leptin on skeletal muscle tissue.

Insulin-like vs. non-insulin-like stimulation of glucose metabolism by vanadium, tungsten, and selenium compounds in rat muscle.

The direct impact of vanadate, tungstate, selenate, and selenite on glucose metabolism of isolated rat soleus muscle was investigated. All compounds stimulated glucose transport, but only vanadate exerted an insulin-like effect on glycogen synthesis (mumol glucose into glycogen*g-1*h-1: control 1.43 +/- 0.11 vs. 1 mmol/l vanadate, 2.08 +/- 0.11, p < 0.0001), which was more distinct in the presence of 1 mmol/l H2O2 (control, 1.44 +/- 0.13 vs. 1 mmol/l vanadate, 3.49 +/- 0.12, p < 0.001). Glucose handling of muscles exposed to tungstate, selenate, or selenite resembled that of hypoxic muscle, i.e. the induced rise in glucose uptake was inhibited by dantrolene and associated with high rates of glycolysis and rapid glycogen depletion (glycogen content after incubation, mumol glucosyl units/g: control, 16.2 +/- 0.7 vs. hypoxia, 2.7 +/- 0.5, p < 0.0001; control, 17.0 +/- 0.5 vs. 100 mmol/l tungstate, 5.5 +/- 0.4, p < 0.001; control, 16.2 +/- 0.7 vs. 100 mmol/l selenate, 1.5 +/- 0.3, and vs. 300 mumol/l selenite, 1.7 +/- 0.3, p < 0.0001 each). The results suggest that vanadate (and more pronounced it's peroxides) exerts true insulin-like action on isolated muscle glucose metabolism, whereas tungsten and selenium salts trigger glucose transport in association with a catabolic response, which may represent an unspecific response to toxic/osmotic stress.