Abstract
Preparations of synaptosomes isolated in sucrose or in Na+-rich media were compared with respect to internal pH (pH1), internal Ca2+ concentration ([Ca2+]i), membrane potential and45Ca2+ uptake due to K+ depolarization and Na+/Ca2+ exchange. We found that synaptosomes isolated in sucrose media have a pHi of 6.77±0.04 and a [Ca2+]i of about 260 nM, whereas synaptosomes isolated in Na+-rich ionic media have a pHi of 6.96±0.07 and a [Ca2+]i of 463 nM, but both types of preparations have similar membrane potentials of about −50 mV when placed in choline media. The sucrose preparation takes up Ca2+ only by voltage sensitive calcium channels (VSCC'S) when K+-depolarized, while the Na+-rich synaptosomes take up45Ca2+ both by VSCC'S and by Na+/Ca2+ exchange. The amiloride derivative 2′, 4′ dimethylbenzamil (DMB), at 30 μM, inhibits both mechanisms of Ca2+ influx, but 5-(N-4-chlorobenzyl)-2′, 4′ dimethylbenzamil (CBZ-DMB), at 30 μM, inhibits the Ca2+ uptake by VSCC'S, but not by Na+/Ca2+ exchange. Thus, DMB and CBZ-DMB permit distinguishing between Ca2+ flux through channels and through Na+/Ca2+ exchange. We point out that the different properties of the two types of synaptosomes studied account for some of the discrepancies in results reported in the literature for studies of Ca2+ fluxes and neurotransmitter release by different types of preparations of synaptosomes.
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Abbreviations
- BCECF:
-
2,7-Biscarboxyethyl-5(6)-carboxyfluorescein
- BCECF/AM:
-
acetoxymethyl ester of BCECF
- [Ca2+]i :
-
Internal free calcium ion concentration
- CBZ-DMB:
-
5-(N-4-chlorobenzyl)-2′,4′-dimethylbenzamil
- DMB:
-
2′, 4′-dimethylbenzamil
- DMSO:
-
dimethyl sulfoxide
- Indo-1/AM:
-
acetoxymethyl ester of Indo-1
- MES:
-
2-|N-Morpholino|ethanesulfonic acid
- NMG:
-
N-methyl-D-glucamine
- pHi :
-
internal pH
- TPP+ :
-
tetraphenylphosphonium
- ΔΨp :
-
plasma membrane potential
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Bandeira-Duarte, C., Carvalho, C.A.M., Cragoe, E.J. et al. Influence of isolation media on synaptosomal properties: Intracellular pH, pCa, and Ca2+ uptake. Neurochem Res 15, 313–320 (1990). https://doi.org/10.1007/BF00968678
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DOI: https://doi.org/10.1007/BF00968678