Evidence for a Serotonin Uptake System in Isolated Bovine Pinealocyte Suspensions

Serotonin uptake was investigated in bovine pinealocyte suspensions. Pinealocytes were disaggregated from calf pineal glands by enzymatic treatment with papain followed by mechanical disruption of the tissue fragments. Cells were resuspended in Hank’s balanced salt solution which contained 12.5 µM nialamide to inhibit monoamine oxidase activity present in the pinealocytes. The uptake of radiolabeled serotonin as a function of concentration exhibited saturation phenomena when high (10 µM-5 mM) concentrations of serotonin were used. Estimates of the fraction of serotonin bound to the cell membrane were attempted by three methods: (1) lysing cells by exposing them to a hypotonic medium after incorporation of radiolabeled serotonin in standard medium; (2) exposing cells to media of increasing osmolarity in the presence of radiolabeled serotonin; and (3) displacing radiolabeled serotonin from nonspecific binding sites on the cell membrane by cold serotonin. Estimates of the fraction of uptake of serotonin contributed by simple diffusion were made by adding an excess of unlabeled serotonin to 0.76µM radiolabeled serotonin to saturate specific carrier sites and nonspecific membrane binding sites. Tissue-to-medium ratios exceeded 1 with all concentrations of serotonin used.