Gary D. Lopaschuk

Glycogen Metabolism in the Aerobic Hypertrophied Rat Heart

BACKGROUND Rates of glycolysis from exogenous glucose are accelerated in hypertrophied hearts. In this study, we determined whether alterations in the metabolism of glycogen, an endogenous storage form of glucose, also occur in hypertrophied hearts. METHODS AND RESULTS Rates of glycolysis ([3H]H2O production) and oxidation ([14C]CO2 production) from exogenous glucose and glycogen were measured in isolated working hearts from control and aortic-banded rats. Hearts in which glycogen was prelabeled with [5-(3)H]- or [U-(14)C]glucose were perfused with buffer containing 11 mmol/L [5-(3)H]- or [U-(14)C]glucose (different from the isotope used to prelabel glycogen), 0.4 mmol/L palmitate, 0.5 mmol/L lactate, and 100 microU/mL insulin. Rates of glycolysis from exogenous glucose were greater (3471+/-114 versus 2665+/-194 nmol glucose x min(-1) x g dry wt(-1), P<.05, n=4 to 6, mean+/-SEM) and rates of exogenous glucose oxidation (445+/-36 versus 619+/-16 nmol glucose x min(-1) x g dry wt(-1), P<.05, n=4 to 6) were lower in hypertrophied hearts than in control hearts. Rates of glycolysis and oxidation from glycogen were not different between hypertrophied and control hearts. A greater proportion of glycogen was oxidized (80% to 100%) than the proportion of exogenous glucose oxidized (13% to 24%) in both groups. Additionally, 10.5+/-1.4 and 12.3+/-1.0 micromol/g dry wt of glycogen was synthesized in hypertrophied and control hearts, respectively, indicating that simultaneous synthesis and degradation (ie, glycogen turnover) occurred in both groups. CONCLUSIONS Thus, aerobic myocardial glycogen metabolism in the hypertrophied heart is similar to that observed in the normal heart even though exogenous glucose metabolism is altered in the hypertrophied heart.

Propionyl L-carnitine improvement of hypertrophied rat heart function is associated with an increase in cardiac efficiency.

Although propionyl L-carnitine improves contractile function of hypertrophied rat hearts, the mechanism(s) by which it does this are not known. One postulated mechanism is that propionyl L-carnitine reverses the alterations in energy metabolism that occur secondary to the carnitine deficiency seen in hypertrophied myocardium. This study determined the effects of chronic propionyl L-carnitine administration on myocardial carnitine content and energy metabolism in hypertrophic hearts from male Wistar Kyoto rats. Pressure-overload hypertrophy was produced by constriction of the abdominal aorta in juvenile rats. Propionyl L-carnitine was administered to the rats via the drinking water for an 8 week period (60 mg.kg(-1).day-1). Myocardial function and metabolic analysis was determined in isolated working hearts obtained from aortic-banded and sham-operated (control) animals at the end of the 8 week study period. Carnitine content was significantly decreased in hypertrophied hearts compared to control hearts, but was normalized by propionyl L-carnitine treatment. Propionyl L-carnitine treatment also prevented the decrease in cardiac work that occurred in hypertrophied hearts compared to control hearts. The primary change in energy substrate use in hypertrophied hearts was a decrease in fatty acid oxidation rates. Glucose and lactate oxidation were similar in control and hypertrophied hearts. While glycolytic rates were slightly higher at moderate workloads, this was not seen at high workloads. Surprisingly, propionyl L-carnitine treatment did not reverse the depression of fatty acid oxidation seen in hypertrophied rat hearts. In fact, a further significant decrease in fatty acid oxidation occurred, such that the contribution of fatty acid oxidation to ATP production decreased from 35 to 26%. Since propionyl L-carnitine treatment increased cardiac work in hypertrophied hearts despite an overall decrease in ATP production rates, an increase in cardiac efficiency was seen. In treated vs. untreated hypertrophied hearts efficiency (cardiac work/ATP produced) increased from 0.23 to 0.40 ml.mm Hg.mumol ATP-1.g dry weight at high workloads. These data suggest that the beneficial effect of propionyl L-carnitine on mechanical function in the hypertrophied heart does not result from a normalization of fatty acid oxidation, but rather from an increase in the efficiency of translating ATP production into cardiac work.

Effect of insulin treatment on long-term diabetes-induced alteration of myocardial function

Six months following the induction of diabetes by streptozotocin (50 mg/kg i.v.) diabetic rats exhibited elevated levels of plasma glucose and glycosylated hemoglobin. Plasma insulin levels were 50% of control and diabetic animals weighed significantly less than control. Using a working heart preparation it was found that (+) and (-) dP/dt and left ventricular pressure development (LVDP) was decreased in hearts from diabetic animals. Insulin treatment (9 U/kg/day s.c. of protamine zinc insulin) for 4 weeks prior to sacrifice restored body weight and plasma insulin to normal. Plasma glucose and glycosylated hemoglobin levels were significantly decreased towards normal in insulin treated diabetic rats. LVDP and (+) dP/dt was also partially returned to normal in insulin treated diabetic rats while (-) dP/dt was completely reversed to normal. Thus, 4 weeks of insulin treatment to rats previously diabetic for 5 months partially or totally reversed the changes produced by diabetes.

Depression of calcium transport in sarcoplasmic reticulum from diabetic rats: Lack of involvement by specific regulatory mediators☆

Cardiac sarcoplasmic reticulum (SR) ATP-dependent Ca2+-uptake was found to be depressed in 4 month streptozotocin-induced diabetic rats. Calmodulin, cAMP-dependent protein kinase and K+ stimulated Ca2+-uptake to similar degrees in SR from both control diabetic rats. Long chain acylcarnitine (7 microM) decreased Ca2+-transport in control rats by 46% but only 26% in diabetic animals. The data suggests that the depression in cardiac SR Ca2+-uptake activity in diabetic rats is non-specific in origin and not a result of alterations in regulation of SR function.

The presence and binding characteristics of calmodulin in microsomal preparations enriched in sarcoplasmic reticulum from rabbit skeletal muscle

ATP-dependent oxalate facilitated calcium transport in sarcoplasmic reticulum (SR) preparations obtained from rabbit vastus lateralis muscle (fast skeletal muscle; Fsr) and soleus (slow skeletal muscle; Ssr) was determined. Addition of exogenous calmodulin did not stimulate calcium transport in either Fsr or Ssr preparations. Fsr and Ssr previously washed in 1 mM EGTA demonstrated a reduced capacity to transport Ca2+; the exogenous addition of calmodulin (0.24 microM) under these conditions, did not restore uptake activity but significantly decreased the steady-state level of Ca2+ uptake. Extracts of skeletal SR prepared by treatment with 0.2 mM EDTA and boiling produced significantly more stimulation of red cell Ca2+ATPase activity than extracts prepared by boiling alone. This stimulation of red cell Ca2+-ATPase was inhibited to a significant extent by 48/80, a known anti-calmodulin agent. Radioimmunoassay revealed that extracts prepared by boiling or EDTA-treatment followed by boiling contained considerable amounts of calmodulin. Washing with 1 mM EGTA, though, did not release any calmodulin from SR. These studies reveal that calmodulin is present in both Fsr and Ssr and can only be removed by harsh treatments. The role of calmodulin in skeletal muscle Ca2+-transport remains to be determined.

Continuous long-term insulin delivery in diabetic rats utilizing implanted osmotic minipumps.

Surgically implanted osmotic minipumps were used to continuously deliver insulin to chemically-induced diabetic rats. Serum glucose levels were maintained within normal limits for 7 days in all diabetic rats implanted with the minipumps. Beyond this time period, serum glucose levels could not be adequately controlled in greater than 50% of the diabetic rats. Seven of 20 diabetic rats originally implanted with osmotic minipumps were well controlled throughout the 31-day study period. This study demonstrates that insulin-filled osmotic minipumps provide no advantage over daily insulin injection in the long term control of diabetes.

Characterization of cardiac microsomal sarcoplasmic reticulum prepared from control and diabetic rats

A method for the preparation of cardiac sarcoplasmic reticulum (SR) from rat heart is described. SR isolated from control and diabetic rats was characterized to determine if differences in calcium transport activity could originate as an artifact of preparation. Electron micrographs of cardiac SR preparations isolated from control and diabetic rats were indistinguishable. The yeild of SR and the mitochondrial membrane contamination were similar in both preparations. These results suggest that depression in diabetic rat cardiac SR microsomal function is not due to nonspecific alterations in the membrane preparations used.