Long lasting action potentials can be triggered in crayfish giant motor neurons by a short depolarizing pulse under different conditions. The concomitant increase in absorbance of the Ca indicator arsenazo III preloaded into the soma, confirms previous observations suggesting that these potential changes are related to a Ca inward current through the soma membrane.The giant motoneurone (MoG) is a large cell, found bilaterally in the crayfish abdominal ganglia. Early work 1 suggested that the cell soma does not show any regenerative action potential in response to a depolarizing pulse, but it is now known that regenerative responses can be recorded under certain conditions. For instance, when the K + conductance of the membrane is reduced (either by TEA 8 or following inactivation by a conditioning depolarisation), a slow action potential (SAP) may be triggered by a short depolarization. This response is most probably due to an influx of Ca 2+ ions, since it depends upon the presence of Ca ~+, but not Na + in the bathing solution, and is blocked by Co 2+ and Mn 2+, but not by TTX 2. In the preparations pretreated with TEA, the SAPs can also be recorded at more hyperpolarized levels than the normal resting potential. TEA is effective either injected into the MoG soma or added to the saline 3. In this paper most of the results were obtained in this way when TEA was added to the solution. We report below that the internal free Ca concentration increases during the SAP, as demonstrated by the change in absorbance of the Ca z+ indicator dye arsenazo lII preloaded into the MoG soma. This observation strengthens previous observations suggesting that SAP is associated .with an influx of Ca 2+ ions. Methods. Experiments were carried out on the giant motor neurons of the crayfish Procambarus clarkii. The ventral chord was dissected and perfused with Van Harreveld's solution containing (in raM): NaC1 195; KC1 5.4; CaC12 13; MgC12 2.6; Tris maleate 10, adjusted to pH 7.2 by NaOH. The 2nd, 3rd and 4th ganglia were desheathed and used for the experiments. The membrane potential was monitored by conventional electrophysiological techniques using a 3 M KC1 electrode. A 2nd electrode filled with an aqueous solution of 1 mM arsenazo III (Sigma grade 1) was also inserted in the MoG soma. The dye was injected by negative current pulses (500 msec, 1 Hz) which hyperpolarized the cell by 30-40 inV. The injection was stopped when the soma appeared just detectably stained as judged by eye. The Arsenazo electrode was then used for passing both the conditioning depolarizing current pulse and the brief (2-5 msec) pulse triggering the SAP. The intracellular calcium transients were monitored by recording the differential absorbance of the dye at 650 and 700 nm as previously described 4,5.Results. Typical responses obtained in TEA-treated preparations are shown in figure 1. The SAP was almost unchanged in Na-free solutions ( fig. 1A). On the contrary, when Ca ++ was omitted from the bathing solution, the SAP was abolished (fig. 1B). These obser...
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