Lability in storage of 3H-dopamine and 3H-norepinephrine in crude synaptosome (P2) and vesicle-associated fractions of rat brain.

Crude synaptosome (P2) fractions prepared from rat striatum and hypothalamus, preloaded with 3H-dopamine (DA) or 3H-norepinephrine (NE), were incubated at 37 degrees C for 5 min. The addition of reserpine at a concentration of 0.1 microM to the striatal synaptosomes substantially depleted 3H-DA to about 45% of control values, but had no effect on 3H-NE. An analogous difference in sensitivity to reserpine, though less pronounced, was observed between 3H-DA and 3H-NE loaded into hypothalamic synaptosomes. Preloaded synaptosome fractions prepared from striatum and hypothalamus were also lysed under hypoosmotic conditions, filtered, and then washed with 130 mM KH2PO4 buffer, pH 7.4, maintained at 0 degrees or 37 degrees C. Washing with 0 degrees C buffer produced no appreciable change in the amount of 3H-DA or 3H-NE retained by the hypoosmotic-resistant subsynaptosomal fractions. Increasing the temperature of the wash buffer to 37 degrees C, however, elicited a volume-dependent depletion of 3H-DA about 2.5-fold higher than that obtained for 3H-NE. Consistent with this finding, the retention of 3H-DA by a crude vesicle fraction prepared from striatum was found to be significantly less than the retention of 3H-NE following 4.5 and 6 min of incubation at 20 degrees C. Thus, in intact synaptosomes, 3H-DA appears to be stored in a form that is more susceptible than 3H-NE to depletion by reserpine, and this effect may be related to differences between the intravesicular storage stability of DA and NE.