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The elemental composition of the presynaptic nerve terminals in rapidly frozen synapses of the cerebellar molecular layer was determined by electron probe x-ray microanalysis and elemental imaging of characteristic x-rays. Elemental imaging of thin freeze-dried cryosections from fresh cerebellar slices frozen within 20 sec of removal from the brain showed normal concentrations of potassium (95 +/- 6 mmol/liter wet tissue +/- SEM) and calcium (0.8 +/- 0.4 mmol/liter) in whole presynaptic terminals, even though mitochondrial and nonmitochondrial sites containing up to 30 mmol of calcium per liter were present elsewhere in the neuropil. Quantitative electron probe analysis of synaptic vesicle clusters and intraterminal mitochondria indicated that their calcium concentrations were 0.4 +/- 0.1 and 1.2 +/- 0.2 mmol/liter, respectively. The low calcium content of presynaptic organelles was confirmed by the absence of detectable deposits in preparations freeze-substituted so as to stabilize calcium content. Similar experiments were carried out on cerebellar slices rapidly frozen after incubation in vitro. The distribution of potassium and calcium in presynaptic terminals of resting and depolarized (55 mM potassium) slices was qualitatively and quantitatively similar to that in freshly excised cortex, although resting slices lacked the few calcium-rich sites that appeared in other areas of the neuropil after stimulation. The calcium concentrations in whole terminals, synaptic vesicles, and mitochondria of resting slices were 1.4 +/- 0.7, 0.7 +/- 0.2, and 0.9 +/- 0.2 mmol/liter, respectively. Thus, amounts of calcium typical of storage organelles in other tissues are not present within cerebellar synaptic vesicles, suggesting that they have a limited role in calcium storage and release.

Distribution of calcium and potassium in presynaptic nerve terminals from cerebellar cortex.