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Intracellular calcium clearance in Purkinje cell somata from rat cerebellar slices
Author(s) -
Fierro Leonardo,
DiPolo Reinaldo,
Llano Isabel
Publication year - 1998
Publication title -
the journal of physiology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.802
H-Index - 240
eISSN - 1469-7793
pISSN - 0022-3751
DOI - 10.1111/j.1469-7793.1998.499bk.x
Subject(s) - cyclopiazonic acid , chemistry , biophysics , extracellular , intracellular , thapsigargin , calcium , endoplasmic reticulum , depolarization , calcium in biology , serca , membrane potential , cytoplasm , biochemistry , biology , atpase , organic chemistry , enzyme
1 The mechanisms governing the return of intracellular calcium (Ca 2+ i ) to baseline levels following depolarization‐evoked [Ca 2+ ] i rises were investigated in Purkinje cell somata using tight‐seal whole‐cell recordings and fura‐2 microfluorometry, for peak [Ca 2+ ] i ranging from 50 nM to 2 μM. 2 Ca 2+ i decay was well fitted by a double exponential with time constants of 0.6 and 3 s. Both time constants were independent of peak [Ca 2+ ] i but the contribution of the faster component increased with [Ca 2+ ] i . 3 Thapsigargin (10 μM) and cyclopiazonic acid (50 μM) prolonged Ca 2+ i decay indicating that sarco‐endoplasmic reticulum Ca 2+ (SERCA) pumps contribute to Purkinje cell Ca 2+ i clearance. 4 A modest participation in clearance was found for the plasma membrane Ca 2+ (PMCA) pumps using 5,6‐succinimidyl carboxyeosin (40 μM). 5 The Na + ‐Ca 2+ exchanger also contributed to the clearance process, since replacement of extracellular Na + by Li + slowed Ca 2+ i decay. 6 Carbonyl cyanide m ‐chlorophenylhydrazone (CCCP, 2 μM) and rotenone (10 μM) increased [Ca 2+ ] i and elicited large inward currents at −60 mV. Both effects were also obtained with CCCP in the absence of external Ca 2+ , suggesting that mitochondrial Ca 2+ uptake uncouplers release Ca 2+ from intracellular stores and may alter the membrane permeability to Ca 2+ . These effects were irreversible and impeded tests on the role of mitochondria in Ca 2+ i clearance. 7 The relative contribution of the clearance systems characterized in this study varied as a function of [Ca 2+ ] i . At 0.5 μM Ca 2+ i , SERCA pumps, PMCA pumps and the Na + ‐Ca 2+ exchanger contribute equally to removal and account for 78 % of the process. Only 45 % of the removal at 2 μM Ca 2+ i can be explained by these systems. In this high [Ca 2+ ] i range the major contribution is that of SERCA pumps (21 %) and of the Na + ‐Ca 2+ exchanger (18 %), whereas the contribution of PMCA pumps is only 6 %.