Energy dispersive x-ray fluorescence and atomic absorption spectroscopy were used to determine the concentrations of Mg, Ca, Mn, Fe, Zn, and Cu in primary cultures of astroglial cells from chick embryo cortex in chemically defined serum-free growth medium. The intracellular volume of cultured glia was determined to be 8.34 microliter/mg protein. Intracellular Mn, Fe, Zn, and Cu in these cells were ca. 10-200 microM, or 20-200 times the concentrations in the growth medium. Mg2+ was 7 mM in glial cells, only four-fold higher than in growth medium. Glutamine synthetase (GS), compartmentalized in glia, catalyzes a key step in the metabolism of neurotransmitter L-glutamate as part of the glutamate/glutamine cycle between neurons and glia. Hormones (insulin, hydrocortisone, and cAMP) added to growth medium differentially altered the activity of GS and the intracellular level of Mn(II), but not Mg(II). These findings suggest the possibility that glutamine synthetase activity could be regulated in brain by the intracellular levels of Mn(II) or the ratio of Mn(II)/Mg(II), which may in turn be controlled indirectly by means of transport processes that respond to hormones or secondary metabolic signals.