The GABAergic innervation of the goldfish pituitary was studied at the light and electron microscope levels by means of radioautography after in vitro incubation in tritiated gamma-aminobutyric acid (GABA) and immunocytochemistry using antibodies against GABA. Following incubation of pituitary fragments in a medium containing tritiated GABA, a selective uptake of the tracer was observed within the digitations of the neurohypophysis. Silver grain clusters were also observed in the adenohypophyseal tissue. At the electron microscope level, this uptake was found to correspond to nerve endings containing small clear and dense-core vesicles. These labeled profiles were located mainly in neurohypophyseal digitations in close apposition with the basement membrane separating the neurohypophysis from the adenohypophysis. However, they were also encountered in direct contact with most adenohypophyseal cell types in the different lobes. These results were confirmed by immunocytochemical data demonstrating the presence of numerous GABA immunoreactive fibers in both anterior and neurointermediate lobes. They were found either in the digitations of the neurohypophysis or in the adenohypophysis in direct contact with the glandular cells with a distribution and an ultrastructural aspect similar to those observed by radioautography. These data demonstrate that the pituitary of teleosts receives a massive GABAergic innervation. Although physiological data providing a functional significance for such an innervation are lacking, the present study suggests that, as already documented in mammals, GABA may be involved in the neuroendocrine regulation of pituitary functions in teleosts.

This work investigated the action of neuropeptide Y (NPY) on thein vitro pituitary release of the maturing gonadotropic hormone (GtH) of the rainbow trout using a perifusion system employing trout balanced salt solution (pH 7.5) at 15°C and a 12.5 ml/h flow rate. In vitellogenic females a 20 minutes NPY application (10(-7) M) induced a 20-30% decrease in GtH secretion. Removal of NPY was followed by a rebound in GTH secretion. On the contrary, in ovulated females, NPY (15 minutes, 10(-7) M) directly stimulated GTH secretion. The greatest stimulation was obtained the day of ovulation where the stimulatory effect of NPY was similar to those induced by s.GnRH in the same conditions, reaching 400% of the basal GTH level. In vitellogenic females treated with 1-4-6 androstadien 3-7 dione, an inhibitor of aromatase activity, the pituitary response to NPY was similar to that obtained in ovulated females. Thus thein vitro action of NPY might depend on thein vivo steroidogenic environment.

To investigate a possible effect of osmotic pressure on prolactin (PRL) release in rainbow trout, we developed a technique for in vitro perifusion of trout pituitaries. Changes in osmotic pressure similar to those observed in fish plasma during transfer experiments did not induce significant modifications of PRL release. In contrast, high-amplitude variation of osmotic pressure resulted in clear modifications of PRL secretion: hyperosmotic medium caused a reduction in PRL release, while infusion of hyposmotic medium induced a transitory increase in PRL release. By using different concentrations of mannitol, we found that the modifications of prolactin secretion could not be ascribed to alterations of the ionic composition of the medium but actually resulted from variations in the osmotic pressure of the incubation medium. In further experiments osmotic pressure was decreased from 300 to 220 mOsm/kg or from 400 to 300 mOsm/kg; a similar transitory increase in PRL release was observed. Measurement of gonadotropin (GtH) in the perifusion effluent medium showed that PRL and GtH secretion followed similar patterns. Thus, our results suggest a possible mechanical effect of wide changes in osmotic pressure on pituitary cell membranes. These data indicate that the rainbow trout differs notably from nonsalmonid teleost species thus far studied in the lack of sensitivity of its PRL cells to osmotic pressure.

Hypothalamic control of prolactin (PRL) release in immature rainbow troutSalmo gairdneri was investigated using anin vitro perifusion system of the rostral pars distalis. Hypothalamic extract of trout induced a dose-dependent stimulation of PRL release. A similar effect was observed when infusing the medium from a 24h static incubation of the hypothalamus. Extracts from different control tissues (muscle, liver, gut) did not changein vitro release, thus confirming the specificity of this stimulatory effect. Hypothalamic extract from adult male rat, known to contain PRL release inhibiting factors, stimulatedin vitro PRL secretion in rainbow trout. This suggests that PRL cells are predominantly influenced by PRL releasing factors. Measurement of TRH and serotonin content in trout hypothalamus indicated consistent physiological levels of these two factors. HPLC studies of hypothalamic extract showed that immunoreactive - TRH eluted at the same place as labelled TRH standard. Moreover, pizotifen, a serotonin antagonist, partially inhibited the stimulation observed with trout hypothalamic extract. These results suggest that, in immature rainbow trout, PRL release is under stimulatory hypothalamic control and that serotonin and probably TRH play a major role in this control.