GLUTAMINE CYCLE ENZYMES IN THE CRAYFISH GIANT NERVE-FIBER - IMPLICATIONS FOR AXON-TO-GLIA SIGNALING

Two of the key enzymes involved in glutamate metabolism, glutaminase a nd glutamine synthetase, were quantitatively localized to axons and gl ia of the crayfish giant nerve fiber by immunocytochemistry and electr on microscopy of antibody-linked gold microspheres. In Western blots, rabbit antisera for glutamine synthetase and glutaminase specifically recognized crayfish polypeptides corresponding approximately in size t o subunits of purified mammalian brain enzymes. Glutamine synthetase i mmunoreactivity was found to be 11 times greater in the adaxonal glial cells than in the axon. Glutaminase immunoreactivity was found in som ewhat greater concentration (2.5:1) in glia as compared to axoplasm. G lutamate immunoreactivity also was evaluated and found to be present i n high concentration in both glia and axons, as might be expected for an important substrate of cellular metabolism. Using radiolabeled subs trates it was demonstrated that glutamine and glutamate were interconv erted by the native enzymes in the intact crayfish giant nerve fiber a nd that the formation of glutamine from glutamate occurred in the axop lasm-free nerve fiber, the cellular component of which is primarily pe riaxonal glia. The results of this investigation provide immunocytoche mical and metabolic evidence consistent with an intercellular glutamin e cycle that modulates the concentration of periaxonal glutamate and g lutamine in a manner similar to that described for perisynaptic region s of the vertebrate central nervous system. These findings further cor roborate previous electrophysiological evidence that glutamate serves as the axon-to-glial cell neurochemical signal that activates glial ce ll mechanisms responsible for periaxonal ion homeostasis. (C) 1995 Wil ey-Liss, Inc.