High endogenous calcium buffering in Purkinje cells from rat cerebellar slices

1. The ability of Purkinje cells to rapidly buffer depolarization-evoked intracellular calcium changes (delta [Ca2+]i) was estimated by titrating the endogenous buffer against incremental concentrations of the Ca(2+)-sensitive dye fura-2. 2. In cells from 15-day-old rats, pulse-evoked delta [Ca2+]i were stable during the loading with 0.5 mM fura-2 through the patch pipette. In cells from 6-day-old rats, delta [Ca2+]i decreased by approximately 50% during equivalent experiments. This decrease was not related to changes in Ca2+ influx, since the integral of the Ca2+ currents remained constant throughout the recording. 3. Experiments with high fura-2 concentrations (1.75-3.5 mM) were performed in order to obtain for each cell the curve relating delta [Ca2+]i to fura-2 concentration. From this relationship, values for the Ca2+ binding ratio (the ratio of buffer-bound Ca2+ changes over free Ca2+ changes) were calculated. 4. In Purkinje cells from 15-day-old rats, the Ca2+ binding ratio was approximately 2000, an order of magnitude larger than that of other neurones and neuroendocrine cells studied to date. This Ca2+ binding ratio was significantly smaller (approximately 900) in Purkinje cells from 6-day-old rats. 5. We propose that the large Ca2+ binding ratio of Purkinje cells is related to the presence of large concentrations of Ca2+ binding proteins and that these cells regulate their ability to handle Ca2+ loads during development through changes in the concentration of Ca2+ binding proteins.