Abstract. A superfusion model for isolated ovarian cells was developed and characterized in detail. Granulosa cells isolated from pre-ovulatory rat ovarian follicles were placed in superfusion (perifusion) chambers with a volume of 125 μl. Culture medium was pumped through the chambers, collected in 20-min fractions of 600 μl and analysed for cAMP and steroids. Viability was confirmed by morphological examination. The use of polycarbonate membranes to retain the cells in the chambers was abandoned since the membranes caused severe cell damage. The temporal relationships between gonadotropic stimuli and the release of cyclic 3':5'-adenosine monophosphate (cAMP) and steroids was investigated. Within 10 min FSH elicited transient increase in the release of cAMP and progesterone but had no effect on testosterone or estradiol-17β release. Amplitude and duration of the response in cAMP and progesterone release were correlated to concentration and length of the FSH pulse when these parameters were varied within the ranges 1–100 μg/l and 30–270 min, respectively. Compared with the cAMP response, the progesterone response peaked up to 30 min later and lasted 1 to 2 h longer but could not be extended to more than approximately 6 h, not even with longer FSH pulses. These results could indicate a development of desensitization.
Carl Johanson and Viktor Johanson
Viktor Johanson, Torsten Ofverholm and Lars E. Ericson
Abstract. A technique for in vivo infusion in the superior thyroid artery in rats and mice was developed and evaluated. The influx catheter is inserted in retrograde direction into the superior carotid artery. The infusate mixed with blood is directed exclusively to the thyroid lobe via the superior thyroid artery. The thyroid isthmus is divided and the other lobe serves as a control. Thyroid ultrastructure was unaltered after infusion for at least 4 h and the follicle cells displayed a normal morphological response to TSH. Electron microscopical autoradiography (125I, [3H]leucine) was performed using 20–80 times less label as compared with iv administration. Infusion of forskolin, a stimulator of adenylate cyclase, increased the intrathyroidal cyclic AMP levels about 10-fold. Infusion of the ionophore monensin yielded typical dilations of Golgi cisternae as well as reduced secretion of newly synthesized protein into the follicle lumen. The arterial infusion technique developed is useful when in vitro methods or systemic administration of sustances are unsuitable. The technique permits selective administration of small amounts of experimental substances to the thyroid in high concentrations.