A new method for quantitating intracellular transport: application to the thyroid hormone 3,5,3'-triiodothyronine

After entering cells from plasma, molecules must permeate through the cytoplasm before they can be metabolized or excreted. If sufficiently slow, cytoplasmic transport may determine the overall rate of cellular elimination at steady state. Cytoplasmic transport of amphipathic molecules should be particularly slow because of extensive binding to intracellular membranes and proteins. Traditional transport models assume that molecules become instantly available for metabolism and canalicular excretion after entering the cell and thus cannot be used to assess cytoplasmic transport. We therefore extended the traditional multiple-indicator dilution (MID) method of Goresky to explicitly incorporate cytoplasmic transport and used the resulting model to estimate the rate constant for cytoplasmic transport of the amphipathic thyroid hormone 3,5,3'-triiodothyronine (T3). We chose T3 because control studies indicated that it is neither metabolized nor excreted during the brief period of an MID experiment (40-90 s). The traditional MID model was unable to account for the data unless we postulated rapid metabolism or excretion of T3. In contrast, the new diffusion MID model fit the data closely without this false assumption and gave values for the influx and efflux rate constants that agreed with previously published data. The half-time for equilibration of T3 across the cytoplasm of the hepatocyte averaged 50 s. This corresponds to an effective cytoplasmic diffusion constant of 3.1 x 10(-8) cm2/s, which is > 100 times slower than expected for free T3 in water. Our results indicate that cytoplasmic transport of this model amphipathic compound is much slower than membrane transport.(ABSTRACT TRUNCATED AT 250 WORDS)