Characterization of sodium-dependent, high-affinity serotonin uptake in rat spinal cord synaptosomes.

ABSTRACT

Synaptosomal accumulation of [3H]serotonin was used to determine if the rat spinal cord possesses a high-affinity neuronal uptake system for serotonin. Two temperature-dependent accumulation processes were found, one sodium-dependent, the second sodium-independent. Sodium-dependent [3H]serotonin accumulation was linear with sodium concentrations up to 143 mM, was associated with the purified synaptosomal fraction (P2B), and decreased 76% by osmotic lysis, 88% by sonication, and 96% by 0.1% Triton X-100. Drug inhibition studies demonstrated fluoxetine to be the most potent inhibitor of this system (IC50 0.075 microM) while desipramine (IC50 0.43 microM) and nomifensine (IC50 0.95 microM) were less potent. Kinetic analysis revealed that sodium-dependent accumulation in purified synaptosomes was saturable at low [3H]serotonin concentrations (Ku = 50 nM, Vmax = 4 pmol/mg protein/min). Sodium-independent [3H]5-HT accumulation was substantially less sensitive to fluoxetine, desipramine and nomifensine. While sodium-independent accumulation was not significantly affected by osmotic lysis, it was markedly increased by prior sonication of tissue. Also, in contrast to sodium-dependent accumulation, sodium-independent accumulation was evenly distributed in all tissue fractions, and was not saturable at low [3H]serotonin concentrations. It is concluded that sodium-dependent [3H]serotonin accumulation reflects uptake into spinal serotonergic nerve terminals while sodium-independent accumulation probably reflects a temperature-sensitive binding to membrane fragments. Comparison to brain uptake of serotonin and the necessity for using 37 degrees C sodium-free blanks rather than 0 degree C blanks in spinal cord homogenates is discussed.