Spontaneous and evoked release of acetylcholine and a cholinergic false transmitter from brain slices: Comparison to true and false transmitter in subcellular stores

Abstract

Rat cerebral cortex slices were incubated with [ 14Ccholine and [ 3Hhomocholine to allow synthesis of [ 14Cacetylcholine and [ 3Hacetylhomocholine; release of the acetylated compounds was tested in the presence of eserine and hemicholinium-3. Both [ 14Cacetylcholine and [ 3Hacetylhomocholine were released spontaneously by a Ca 2+-independent mechanism; exposure of the tissue to a high K + medium resulted in a Ca 2+-dependent increase in [ 14Cacetylcholine release (30-fold) and in [ 3Hacetylhomocholine release (5 to 7-fold). Thus, spontaneous and K +-evoked transmitter release could be distinguished on the basis of their molar ratios of true to false transmitters and these ratios were compared to the molar ratios of the transmitters in subcellular fractions prepared from the incubated tissue. The molar ratio of acetylcholine to acetylhomocholine released from the tissue spontaneously differed from the ratio in subcellular fractions containing occluded transmitters. The molar ratio of acetylcholine to acetylhomocholine released by K + differed from the ratio in a fraction (H) containing occluded transmitters but was similar to the ratio in the fraction (D) containing monodisperse synaptic vesicles and the fraction (0) containing the majority of a soluble cytoplasmic marker. Comparison of the transmitter contents of subcellular fractions from unstimulated and K +-stimulated tissue showed that the three fractions (D, H and O) lost equal proportions of both transmitters as a result of K + stimulation. It is concluded that acetylhomocholine may be released from brain slices both spontaneously and in response to stimulation via mechanism similar to those that release acetylcholine, but that there must be some differences in specificity of the acetylcholine storage and/or release processes. The results also indicate that spontaneous transmitter release originates from extravesicular stores; the results are consistent with a vesicular site of origin of evoked transmitter release but do not distinguish between nerve terminal vesicles and cytoplasm as the immediate source of evoked transmitter release.