[14C]Glucose Metabolism in Sympathetic Ganglia of Chicken Embryos and in Primary Cultures of Neurons and of Other Cells from These Ganglia

Metabolism of [1-14C]glucose and [6-14C]glucose was measured in sympathetic ganglia excised from chicken embryos 12-16 days old and in primary cultures of neurons or nonneurons prepared from these ganglia. Some metabolic rates tended to change with the tissue/medium ratio, so this variable had to be controlled. Less C-6 than C-1 od glucose was put out in CO2 by all three types of preparations, indicating operation of the hexosemonophosphate shunt. The C-6/C-1 ratio was greater for the neuronal cultures and for intact ganglia than for the nonneuronal cultures. The C-6/C-1 ratio for the neurons increased with the amount of tissue added to a given volume of incubation medium, in agreement with previous experiments on embryonic dorsal root ganglia (Larrabee, 1978). Per unit of protein, the output of C-1 of glucose in CO2 was higher in both the neuronal and the nonneural cultures than in intact ganglia, whereas that of C-6 was higher in the neuronal cultures and lower in the nonneuronal ones than in the ganglia. The rates of release in lactate of C-1 and C-6 of glucose were 3-5 times higher from both types of cultures than from intact ganglia. The average rates of incorporation of C-1 and C-6 of glucose into tissue constituents were lower in the cultures than in intact ganglia, significantly so for incorporation of C-6 in the nonneuronal cultures.     

Ontogeny of Glucose and Lipid Metabolism in Dorsal Root Ganglia of Chickens.: Similarities and Contrasts with Sympathetic Ganglia

Dorsal root ganglia, excised from the lumbar roots of the sciatic nerve of white Leghorn chicken embryos 6-13 days of age, were incubated usually for 5 h, at 36 degrees C in 20 microliters of a bicarbonate-buffered physiological salt solution containing 5.5 mM glucose. [U-14C]Glucose, [1-14C]glucose, [6-14C]glucose, or [5-3H]uridine was also added. Lipid synthesis and lactate output were measured by incorporation of 3H from [5-3H]uridine. Glucose uptake and labeled lactate output declined rapidly from 6 to 8-9 days of age, more slowly thereafter. Synthesis of lipids was relatively constant throughout the ages studied, without the increased rate at intermediate ages seen previously in sympathetic ganglia of the same species. RNA synthesis declined progressively throughout the ages studied. The output of C-6 of glucose to CO2 was about the same at all ages, whereas that of C-1 declined rapidly from 6 to 7 days of age and then more slowly, but always remained higher than that of C-6 and thus indicated that much glucose was metabolized via the hexosemonophosphate shunt.     

Ontogeny of Glucose Metabolism in Sympathetic Ganglia of Chickens. Changes in the Carbon Fluxes to CO2, Lactate, and Tissue Constituents from 8 to 19 Days of Embryonic Age

Chains of sympathetic ganglia were excised from the lumbar region of white Leghorn chicken embryos, 8-19 days of age. The chains were incubated for 5 h at 36 degrees C in a bicarbonate-buffered physiological salt solution containing 5.55 mM unlabeled glucose and tracer amounts of glucose labeled either uniformly or at carbon-1 or carbon-6. Glucose uptake and labeled lactate output were both highest in ganglia from the youngest embryos studied and declined progressively with increasing age. The output of labeled CO2 rose to a peak rate at an incubation age of 10-12 days in the presence of either [U-14C]glucose or [1(-14)C]glucose, but changed relatively little with age in the presence of [6(-14)C]glucose. The incorporation of 14C into tissue constituents was fastest at 10-12 days with all three labeled glucoses. It is concluded that the hexosemonophosphate shunt is most active at an incubation age of 10-12 days, after glycolysis has greatly slowed. The literature on morphological and biochemical changes in the sympathetic ganglia during development is briefly reviewed and discussed in relation to the observed metabolic changes. The early high glycolytic rate may be related to the normal developmental delay in vascularization of the sympathetic chains.     

Ontogeny of Glucose Metabolism in Sympathetic Ganglia of Chickens. Concurrence of Maximum Rates in the Hexosemonophosphate Shunt and in Synthesis of Lipids but Not of Ribonucleic Acid

Chains of sympathetic ganglia were excised from the lumbar region of white Leghorn chicken embryos, 8-19 days of age, and incubated, usually for 5 h, at 36 degrees C in a bicarbonate-buffered physiological salt solution containing [U-14C]glucose, [1-14C]glucose, [6-14C]glucose, or [5-3H]uridine. Lipid synthesis was measured by the incorporation of 14C into lipids, and RNA synthesis by the accumulation of 3H into macromolecules. The ratio of 14C put out in CO2 during the second hour of incubation in the presence of [1-14C]glucose to that with [6-14C]glucose was used as an index of activity in the hexosemonophosphate shunt (HMS). Both the rate of lipid synthesis and activity in the HMS reached well-defined maxima at about 11 days of embryonic age. There was no evidence of a similar rise and fall of RNA synthesis during the ages studied. Estimates of the rate of NADPH production by the HMS at near-peak lipid synthesis varied over a twofold range that included the rate needed for the observed lipid synthesis. The results thus support, quantitatively as well as qualitatively, the supposition that the HMS is accelerated during development to sustain lipid synthesis.     

Lactate Uptake and Release in the Presence of Glucose by Sympathetic Ganglia of Chicken Embryos and by Neuronal and Nonneuronal Cultures Prepared from These Ganglia

Abstract: Uptake and output of lactate were measured in lumbar sympathetic chains excised from embryos of white leghorn chickens, 14–15 days old. The chains, typically containing 30–40 μg of protein, were incubated in Eagle's minimum essential medium containing bicarbonate buffer, 6–17 mM glucose, various concentrations of lactate, and either [U-¹⁴C]lactate, [1-¹⁴C]glucose, or [6-¹⁴C]glucose. The average rate of uptake of labeled lactate was measured with incubations of 5–6 h, starting with various external lactate concentrations. From these data the instantaneous relation between lactate uptake rate and concentration was deduced with a simple computerized model. The instantaneous uptake rate increased with the concentration according to a relation that fit the Michaelis-Menten equation, with V_max = 360 μmol/g protein/h and K_m = 4.8 mM. Substantial fractions of the lactate carbon were recovered from tissue constituents and in several nonvolatile products in the medium, as well as in CO₂. Glucose uptake averaged about 108 μmol/g protein/h and did not vary greatly with external lactate concentration, although the metabolic partitioning of glucose carbon was considerably affected. Regardless of initial concentration, the lactate concentration in the medium tended to change towards approximately 0.6 mM, showing that uptake equaled output at this level, with rates at about 40 μmol/g protein/h. With the steady-state concentration of 0.6 mM lactate, about 20% of the glucose carbon was shunted out into the medium before it was reabsorbed and metabolized into various products. Lactate uptakes by neuronal and nonneuronal cultures prepared from the ganglia did not differ consistently from one another or from uptake by undissociated ganglia. The neuronal cultures tended to oxidize a greater fraction of the consumed lactate to CO₂ and to convert a smaller fraction of the lactate to products in the medium than did the nonneuronal cultures. Computer modeling, using known parameters for blood-brain transport of lactate in the adult rat and data on uptake by the ganglia, suggests that lactate may supply substantial fuel to the brain, even in the presence of abundant glucose, when the lactate concentration in the blood is raised to levels commonly observed in exercising humans, such as 10–20 mM. This is in agreement with the findings of several investigators in hypoglycemic humans and in animals with intermediate blood lactate concentrations.     

Partitioning of CO2 Production Between Glucose and Lactate in Excised Sympathetic Ganglia, with Implications for Brain

Abstract: Chains of lumbar sympathetic ganglia from 15-day-old chicken embryos were incubated for 4 h at 36°C in a bicarbonate-buffered salt solution equilibrated with 5% CO₂-95% O₂. Glucose (1–10 m M ), lactate (1–10 m M ), [U-¹⁴C]glucose, [1-¹⁴C]glucose, [6-¹⁴C]glucose, and [U-¹⁴C]lactate were added as needed. ¹⁴CO₂ output was measured continuously by counting the radioactivity in gas that had passed through the incubation chamber. Lactate reduced the output of CO₂ from [U-¹⁴C]glucose, and glucose reduced that from [U-¹⁴C]lactate. When using uniformly labeled substrates in the presence of 5.5 m M glucose, the output of CO₂ from lactate exceeded that from glucose when the lactate concentration was >2 m M . The combined outputs at each concentration tested were greater than those from either substrate alone. The ¹⁴CO₂ output from [1-¹⁴C]glucose always exceeded that from [6-¹⁴C]glucose, indicating activity of the hexose monophosphate shunt. Lactate reduced both of these outputs, with the maximum difference between them during incubation remaining constant as the lactate concentration was increased, suggesting that lactate may not affect the shunt. Modeling revealed many details of lactate metabolism as a function of its concentration. Addition of a blood-brain barrier to the model suggested that lactate can be a significant metabolite for brain during hyperlactemia, especially at the high levels reached physiologically during exercise.