Neuronal galvanotropism is independent of external Ca super(2+) entry or internal Ca super(2+) gradients

The mechanism by which growing neurites sense and respond to small applied electrical fields is not known, but there is some evidence that the entry of Ca super(2+) from the external medium, with the subsequent formation of intracellular Ca super(2+) gradients, is important in this process. We have employed two approaches to test this idea. Xenopus spinal neurites were exposed to electrical fields in a culture medium in which Ca super(2+) was chelated to very low levels compared to the normal extracellular concentration of 2 mM. In other experiments, loading the neurites with the calcium buffer, 1,2-bis(o-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid (BAPTA), disrupted the putative internal Ca super(2+) gradients, and the effects on the electrical response were determined. Fields of 100 mV/mm were applied for 12 h, and no difference was detected in the cathodal turning response between the treated neurites and the untreated controls. Using the Differential Growth Index (DGI), an asymmetry index, to quantitate the turning response, we recorded DGIs of -0.64, -0.65, and -0.62 for control cells, cells in Ca super(2+)-free medium, and cells preloaded with BAPTA, respectively. Furthermore, we detected an increase in neurite length for those neurons cultured in Ca super(2+)-free medium; they were 1.5-1.7 times as long as neurites from neurons cultured in normal Ca super(2+) medium. Likewise, we found that BAPTA-loaded neurites were longer than control neurites. Our data indicate that neuronal galvanotropism is independent of the entry of external Ca super(2+) or of internal Ca super(2+) gradients. Both cell-permeant agonistic and antagonistic analogs of cyclic 3',5'-adenosine monophosphate (cAMP) increased the response to applied electrical fields.