Low-affinity uptake of [3H]choline by rat neurointermediate lobe in vitro

Uptake of [3H]choline by rat neurointermediate lobes in vitro was investigated. The rate of uptake showed saturation with concentration of [3H]choline above 120 microM. Lowering of the incubation temperature from 32 to 4 degrees C, or increasing the concentration of K+ in the incubation medium from 5 to 100 mM, diminished the rate of uptake by 83.8% (SEM 6.9%, n = 3) or 43.1% (SEM 13.5%, n = 3), respectively. Following preloading with [3H]choline, a slow efflux (1% of the content every 20 min) of [3H]radioactivity was observed from the perifused glands. This efflux was enhanced 10-fold by increasing the concentration of K+ in the perifusion medium to 100 mM. Neither the uptake of [3H]choline, nor the subsequent basal or potassium-enhanced efflux of [3H]radioactivity were affected by reducing the concentration of Na+ from 125 to 19 mM, or by including 10 microM hemicholinium-3 in the medium during preincubation and perifusion. Replacing Ca2+ by 0.5 mM EGTA during perifusion resulted in a minor decrease (28%, SEM 7.3%, n = 7) in the potassium-enhanced [3H]radioactivity efflux. This decrease occurred only in one of the two high-potassium periods during the perifusion. In conclusion, uptake of [3H]choline by rat neurointermediate lobes was due to a low-affinity, saturable mechanism, with the efflux of [3H]radioactivity most likely representing the depolarization-facilitated outflow of [3H]choline. Autoradiography of the tissue sections showed this uptake to be localized to both neuronal and glial elements of the neural lobe, in contrast to sparse labelling of pars intermedia.