MECHANISMS OF GLUTAMATE ACTIVATION OF AXON-TO-SCHWANN CELL SIGNALING IN THE SQUID

Membrane potentials from Schwann cells associated with giant axons of the small squid (Alloteuthis and Loliguncula) and the large squid (Loligo) were monitored with glass microelectrodes following 100 Hz/15 s axonal stimulation, or the application of 10(-7) M glutamate and ion substitutions, in the presence or absence of 10(-7) M d-tubocurarine. Glutamate or stimulation caused the membrane of the Schwann cell to depolarize to approximately -32 mV. This was rapidly replaced by a transient hyperpolarization to approximately -55 mV; the potential returning to the resting level (-40 mV) in approximately 7 min. In the presence of d-tubocurarine only the initial depolarization was evident. Nominally zero [Na+]o or treatment with 10(-7) M tetrodotoxin (in normal [Na+]o) blocked the stimulation- and glutamate-induced depolarization while low Cl(o)- hyperpolarized the Schwann cell without effect on the glutamate- or stimulation-induced depolarization. Na(o)+ depletion or pretreatment with tetrodotoxin in normal Na(o)+ did not affect the development of the Schwann cell hyperpolarization. These results do not support the hypothesis that the glutamate-induced depolarization is the trigger leading to the Schwann cell hyperpolarization. Preliminary experiments to test the possibility that inositol phosphate second messenger and an increase in [Ca2+]i are triggered by glutamate receptor activation showed that nominally 0 Ca(o)2+/75 mM Mg(o)2+ only slightly reduced the hyperpolarizing response to stimulation or glutamate while intracellular Bapta (20-30-mu-M) blocked the hyperpolarization but not the depolarization. [H-3]Myoinositol incorporation into axon-Schwann cell plasma membranes was high. Treating the preparation for a period of 1-10 min with the glutamate receptor antagonist, 2-amino-4-phosphonobutyrate (10(-5) M), in the presence of glutamate reduced membrane [H-3]myoinositol content to 45% of control. These observations are consistent with a glutamate-activated Na+ channel for the initial depolarization and an independent inositol phosphate second messenger-Ca2+-dependent triggering event for the development of the Schwann cell hyperpolarization.