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Regulation of Calcium Concentrations in Synaptosomes: α 2 ‐Adrenoceptors Reduce Free Ca 2+ by Closure of N‐Type Ca 2+ Channels
Author(s) -
Xiang JianZhong,
Morton John,
Brammer Michael J.,
Campbell Iain C.
Publication year - 1990
Publication title -
journal of neurochemistry
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.75
H-Index - 229
eISSN - 1471-4159
pISSN - 0022-3042
DOI - 10.1111/j.1471-4159.1990.tb08852.x
Subject(s) - calcium , chemistry , isotopes of calcium , synaptosome , closure (psychology) , biophysics , biology , organic chemistry , economics , market economy
The relationship between intrasynaptosomal total (Ca T ) and free ([Ca 2+ ] i ) calcium and 45 Ca accumulation was studied under physiological and K + ‐depolarised conditions in rat cortical synaptosomes. Under physiological conditions, Ca T (10.7 m M ) was ∼ 10,000 times higher than [(Ca 2+ ] i (118 n M ), showing that there is a large reservoir of sequestered calcium in synaptosomes. 45 Ca accumulation was rapid (initial rate, 3.4 nmol/mg protein/min), substantial (7 nmol/mg protein in 2 min), and depolarisation dependent, and reached equilibrium after 5 min. At equilibrium, only 10% of Ca T was freely exchangeable. This pool was much larger than the free Ca 2+ pool. Ca T , [Ca 2+ ] i , and 45 Ca accumulations were directly related to the Ca 2+ concentration in the buffer, suggesting that [Ca 2+ ] i is not highly conserved but is maintained by simple equilibria between the various pools. Clonidine reduced 45 Ca accumulation in a time‐ and dose‐dependent manner. Maximum inhibition (40% at 100 μM ) occurred at 2 min and the IC 50 was 80 n M The reduction caused by clonidine (1 μM ) reached equilibrium after 5 min, but this equilibrium value was lower than in controls, suggesting that clonidine changes the exchangeable Ca 2+ pool size. The effects of clonidine (1 μM ) on [Ca 2+ ] i (26% reduction) and on 45 Ca accumulation (24% reduction) were most apparent under physiological conditions. However, while it was not dependent on depolarisation, it did not occur in physiological buffer containing low K + concentration (0.1–1 m M ). The inhibitory effect of clonidine on 45 Ca accumulation is receptor mediated as it was antagonised by idazoxan (1 μM ). Under nondepolarised conditions (5 m M K + ), ω ‐conotoxin fraction GVIA ( ω ‐CgTx; 0.5 μM ) caused a 35% reduction in 45 Ca accumulation in a 2‐min incubation, whereas nifedipine (1 μM ) had no significant effect. In contrast, both ω ‐CgTx and nifedipine effectively blocked 45 Ca entry into depolarised synaptosomes. However, the effect of ω ‐CgTx was not additive with that of clonidine, whereas that of nifedipine was. Finally, idazoxan (1 μM ) did not alter the effect caused by nifedipine. These data show (a) that clonidine decreases 45 Ca accumulation by inhibiting N‐ but not L‐type voltage‐sensitive Ca 2+ channels, (b) that specific Ca 2+ channels on synaptosomes display a range of voltage sensitivities, and (c) that L‐type Ca 2+ channels are operational under depolarising conditions.