The distribution of free calcium in transected spinal axons and its modulation by applied electrical fields

Intracellular free-calcium concentration (Ca2+i) was measured in lamprey spinal axons using the fluorescent calcium indicator fura 2. We used both a photomultiplier tube and a video-image processing system to measure the temporal and spatial distributions of Ca2+i in the proximal segments of transected axons. Within 3 min following transection, a spatially graded increase in the Ca2+i was apparent in the last few millimeters of the axons. Superimposed on the initial gradient was a moving front of calcium that progressed up the axon, reaching 1.6 mm from the cut end in 3 hr. The Ca2+i behind the moving front exceeded 10 microM. This movement of Ca2+ was greatly reduced by an externally applied electrical field with the cathode distal to the lesion and was increased by an applied field of the opposite polarity. When axons were transected in Ca2(+)-free medium, no increases in Ca2+i occurred. One d after transection, Ca2+i was at or below the precut levels, except in the distal 250 microns, where it remained slightly elevated. Therefore, axons can survive the high levels of Ca2+i that occur after transection and can reestablish normal Ca2+i levels within 24 hr. Measurements of both the diffusion coefficient and the fluorescence polarization of fura 2 indicate that the dye is not significantly bound to axoplasmic components.