Mapping calcium transients in the dendrites of Purkinje cells from the guinea-pig cerebellum in vitro.
1. A 10 X 10 photodiode array was used to detect stimulation-dependent absorbance changes simultaneously from many positions in the dendrite field of guinea-pig Purkinje cells which had been injected with the calcium indicator Arsenazo III in thin cerebellar slices. Signals from each element of the array were matched to positions on the cells by mapping them onto fluorescence photographs of Lucifer Yellow which had been co-injected into the cells with the Arsenazo III. 2. In response to intrasomatic stimulation the rising phase of the absorbance signals corresponded in time with the calcium spikes recorded with an intracellular electrode. There was no increase in absorbance during bursts of fast sodium spikes. Absorbance signals persisted after the sodium spikes were blocked by tetrodotoxin (TTX). In addition, the signals were largest at 660 nm and small signals of opposite polarity were found at 540 nm. These results indicate that the absorbance signals came from calcium entry into the cell resulting from the turning on of voltage-dependent calcium conductances. 3. In these experiments signals were usually seen all over the dendritic field and were weak or totally absent over the soma. In some cases signals were seen over a more restricted area. With a spatial resolution of 25 microns we were not able to see any evidence for highly localized sites of calcium entry. 4. Sometimes the rising phase of the calcium signals was separated by almost 13 ms in different parts of the dendritic field, too long to be explained by active propagation delay. This suggests that calcium spikes causing these signals can be evoked separately in different regions of the Purkinje cell dendritic field by long-lasting potentials which may reach local threshold at different times. 5. Calcium signals resulting from slow plateau after-potentials and the calcium spikes produced by them were also detected in all locations in the dendritic field. The relative distribution of amplitudes from these plateau signals was different from the distribution of evoked signals during current injection. 6. Climbing fibre synaptic activation produced calcium signals which were distributed over the dendritic arborization, but larger at the main dendritic tree where most of the synaptic contacts are located. 7. Calcium signals were also detected from the dendrites of other neurone types in the in vitro slice preparation. Thus, it is likely that these kind of measurements can be used to analyse the electroresponsiveness of many kinds of neurones in the mammalian brain.
Ross
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WN
Department of Physiology, New York Medical College, Valhalla 10595.
Werman
R
R
D000200:Action Potentials / Q000187:drug effects; D000818:Animals; D001150:Arsenazo III; D001931:Brain Mapping; D002118:Calcium / Q000502:physiology*; D002531:Cerebellum / Q000502:physiology*; D003712:Dendrites / Q000502:physiology*; D006168:Guinea Pigs; D066298:In Vitro Techniques; D011689:Purkinje Cells / Q000502:physiology*; D013779:Tetrodotoxin / Q000494:pharmacology; D013997:Time Factors
D001150:Arsenazo III; D013779:Tetrodotoxin; D002118:Calcium