Effect of Norepinephrine on Glucose Metabolism in Glioblastoma and Neuroblastoma Cells in Cell Culture

Abstract

The addition of norepinephrine to cultured glioblastoma cells results in an inhibition of uptake of radioactivity from D-[2_3H]glucose, D_[1_14C]glucose, D42-14C]glucose, and D- [6-14C]glucose. In addition, if the glioblastoma cells are previously labeled with these substrates, norepinephrine causes an increase in the release of radioactivity. These effects were not observed with cultured neuroblastoma cells. It is suggested that the breakdown of glycogen is activated by norepinephrine as a result of an increase in 3': 5'-cyclic AMP.Several years ago Galambos (1) postulated that, in addition to neuron-neuron synapses, neuron-glia synapses occur. A metabolic relationship existing between neurons and glia was suggested originally by Hyden and Lange (2) on the basis of whole animal studies and more recently by Gilman and Nirenberg (3) on the basis of the finding that the neurotransmitter, norepinephrine, causes an increase in 3': 5'-cyclic AMP (cAMP) in rat glioblastoma cells in cell culture. These findings were substantiated by Clark and Perkins (4), who also found a rapid increase in the concentration of cAMP of epinephrine and histamine, in addition to norepinephrine, in a tumor astrocyte cell-line derived from a primary culture of a human glioblastoma multiforme. It is known from the early work of Rall and Sutherland (5) that glycogenolysis was increased by cAMP due to activation of a protein kinase that converts phosphorylase b to phosphorylase a, the active form of the enzyme.On the basis of these several results, it seemed of interest to examine in cell culture the effect of norepinephrine on glucose incorporation and excretion in glioblastoma and neuroblastoma cells. MATERIALS AND METHODS Cell line and cultureClone C-6, derived from a rat (Wistar strain) astrocytoma induced by N-nitrosomethylurea, and clone C-46, derived from a mouse neuroblastoma, were obtained from Dr. G. Sato (6). Clones C-6 and C-46 were grown in Dulbecco's modified Eagle's (7) medium with 10% fetal-calf serum in Falcon flasks or petri dishes at 370 in an atmosphere of 10% C02-90% air at 100% humidity. Radioactivity experimentsFor the radioactivity experiments, 60-mm petri dishes were incubated with about 3 X 106 cells for 2 days in the above medium. By this time the cells were confluent. The medium was then changed to 2 ml of fresh modified Eagle's medium or Earle's (8) balanced salt solutions containing the radioactive compounds. For the studies on the effect of theophylline, the cells were incubated for 1 hr in the culture medium plus 1 mM of theophylline before the addition of the radioactive medium. Norepinephrine (17 jg/ml) was added where indicated. In the experiments in which cells were previously labeled with radioactive compounds, the cells were incubated for 2 hr in a radioactive medium, washed twice with nonradioactive medium, and then incubated for 30 min in a nonradioactive medium.After incubation in the desired medium the cells were scraped from the petri dish, filtered through a Millipore filter (HAWP 025, HA 0.4...

D‐Glucose Transport in Cultured Cells of Neural Origin: The Membrane as Possible Control Point of Glucose Utilization

“…In agreement with other authors (Passonneau and Crites, 1975) we found much higher glycogen levels in C6 glial cells than in neuroblastoma cells, in which glycogen as source of energy seemed to be unimportant. The occurrence of high glycogen levels within astroglial cell types might support the concept of a nutritional cooperation between glial and neuronal cells in situ, as discussed elsewhere (Newburgh and Rosenberg, 1972). In contrast to neuroblastoma cells, the glycogen content of C6 glioma diminished with increasing cell density and time in culture (Fig.…”

Purification of Glycogen Phosphorylase from Bovine Brain and Immunocytochemical Examination of Rat Glial Primary Cultures Using Monoclonal Antibodies Raised Against This Enzyme

“…T h e functional significance of brain glycogen reserves remains to be elucidated. However, the findings that glycogenolysis is under hormonal control in brain slices (Quach et al, 1978(Quach et al, , 1980Ververken et al, 1982;Magistretti e t al., 1984Magistretti e t al., , 1986, in spinal cord (Woolf, 1987), and in glioma cell lines (Newburgh and Rosenberg, 1972;Drummond et al, 1977; C u m m i n s et al., 1983) support the notion that this metabolic pathway can be modulated by a range of neurohormones and neurotransmitters.…”

On the Role of Calcium Ions in the Regulation of Glycogenolysis in Mouse Brain Cortical Slices

“…The effect of NA on glycogenolysis could be mainly on glial cells rather than on neurons. Indeed, NA induces glycogenolysis in glioblastoma cells, and in astrocytoma cells but not in neuroblastoma cells (Newburgh and Rosenberg, 1972;Opler and Makman, 1972;Passonneau and Crites, 1976); preceptors have been shown to be present in glial cells (Schubert et al, 1976;McMorris, 1977;Brostrom et al, 1979). Moreover, the highly diffuse spatial organization of NA neurons in the cortex (Descarries et al, 1977) can accommodate additional neuron-glia contact, allowing for a noradrenergic control of glial metabolism.…”

“…Brain contains indeed substantial amounts of glycogen phosphorylase and of glycogen synthase, as well as the converting enzymes responsible for rapid activation and inactivation Norman, 1962, 1965;Drummond et al, 1964;Goldberg and 0 Toole, 1969;Drurnrnond and Bellward, 1970). The involvement of cyclic AMP in the control of brain glycogenolysis has been shown in cultured cells (Newburgh and Rosenberg, 1972;Opler and Makman, 1972;Browning et al, 1974;Passonneau and Crites, 1976), in brain slices Makman, 1976, 1977;Quach et al, 1978), and in vivo (Edwards et al, 1974;Folbergrova, 1975a,6;Lust and Passonneau, 1976;Anchors and Garcia-Rill, 1977); however, such a role of cyclic AMP has not been systematically ob-served, in particular with brain slices (Kakiuchi and Rall, 1968;Rall and Gilman, 1970;Edwards et al, 1974). Consequently, it has been suggested that Caz+ may also be important in the regulation of glycogenolysis in nervous tissue (Nahorski et al, 1975;Folbergrova, 1977;Quach et al, 1980), the more so as brain phosphorylase kinase is a calcium-dependent enzyme (Ozawa, 1973).…”