**3. The effect of the day length and exogenous melatonin on the secretion of prolactin under** *in vitro* **conditions**

Studies carried out on a group of lactating sheep under in vivo conditions showed modulating effects of melatonin with respect to the secretion of prolactin. The above mentioned relations demonstrate that melatonin can modulate the seasonal rhythm of the prolactin secretion affecting directly the pituitary gland. The highest concentration of melatonin receptors in the sheep's pituitary gland was identified in the pars tuberalis [36]. This structure is located precisely between the eminentia medialis of the hypothalamus and the distal part of the gland, which enables its mediation in the communication of the brain with the pituitary gland. In the sheep's PT only a MT1 melatonin receptor is found, and surgical separation of the pituitary gland from the hypothalamus does not have any influence on the concentration and location of this receptor and its sensitivity to binding melatonin [37]. Despite the fact that it was demonstrated that the activity of PT secretory cells is seasonal and melatonin-dependent, it was not to confirmed that it is involved in the control of seasonal changes in the sexual activity.

The presence of melatonin receptor limited to the PT of the pituitary gland, and lack of direct effects of melatonin both on the prolactin gene expression and release of prolactin in cultures of lactotroph cells under in vitro conditions [38] suggests that melatonin regulates the seasonal secretion of prolactin by way of a specific compound synthesized in the PT. The presence of such a compound in the PT, with a peptide structure and stimulating secretion of prolactin from lactotroph cells of the pituitary gland, was confirmed in experiments, and the peptide itself was called tuberalin [36,39]. Own studies carried out under in vitro conditions demonstrated that both the day length and administration of exogenous melatonin affect the profile of the prolactin secretion. In vitro incubations of pituitary explants taken from sheep on 30th day of lactation were held in three periods: in the period of lengthening days (March), in the period of shortening days (August) and in the short-day period (November). The pituitaries taken were divided along the sulcus into two halves so that each one contained the adenohypophysis and the neurohypophysis. In vitro incubation was held during 3 hours in the Parker medium at a temperature of 370C. The control group (G1) was incubated in a medium without any hormonal additives, whereas the experimental group (G2) was incubated in a medium with exogenous melatonin. The experiments performed demonstrated that administration of exogenous melatonin in the long-day period caused a decrease of the secretion of prolactin in the pars tuberalis (table 5)

126 Prolactin

Introduction of melatonin implants for sheep rearing lambs in the short-day period did not cause significant changes in the profile of the prolactin secretion. By analyzing the profile of the PRL secretion in both sheep groups, a conclusion can be drawn that in sheep lambed in November the concentration of prolactin in the first control sample drawn was lower by 50% as compared to the control sample drawn in March. In the long-day period the concentration of prolactin in the control group increased, and the secretion of melatonin decreased. In the autumn and winter period, though, as natural conditions set in, the concentration of melatonin increased and the level of prolactin dropped. The extending signal of melatonin observed in the autumn and winter period causes under natural

**3. The effect of the day length and exogenous melatonin on the secretion** 

Studies carried out on a group of lactating sheep under in vivo conditions showed modulating effects of melatonin with respect to the secretion of prolactin. The above mentioned relations demonstrate that melatonin can modulate the seasonal rhythm of the prolactin secretion affecting directly the pituitary gland. The highest concentration of melatonin receptors in the sheep's pituitary gland was identified in the pars tuberalis [36]. This structure is located precisely between the eminentia medialis of the hypothalamus and the distal part of the gland, which enables its mediation in the communication of the brain with the pituitary gland. In the sheep's PT only a MT1 melatonin receptor is found, and surgical separation of the pituitary gland from the hypothalamus does not have any influence on the concentration and location of this receptor and its sensitivity to binding melatonin [37]. Despite the fact that it was demonstrated that the activity of PT secretory cells is seasonal and melatonin-dependent, it was not to confirmed that it is involved in the

The presence of melatonin receptor limited to the PT of the pituitary gland, and lack of direct effects of melatonin both on the prolactin gene expression and release of prolactin in cultures of lactotroph cells under in vitro conditions [38] suggests that melatonin regulates the seasonal secretion of prolactin by way of a specific compound synthesized in the PT. The presence of such a compound in the PT, with a peptide structure and stimulating secretion of prolactin from lactotroph cells of the pituitary gland, was confirmed in experiments, and the peptide itself was called tuberalin [36,39]. Own studies carried out under in vitro conditions demonstrated that both the day length and administration of exogenous melatonin affect the profile of the prolactin secretion. In vitro incubations of pituitary explants taken from sheep on 30th day of lactation were held in three periods: in the period of lengthening days (March), in the period of shortening days (August) and in the short-day period (November). The pituitaries taken were divided along the sulcus into two halves so that each one contained the adenohypophysis and the neurohypophysis. In vitro incubation was held during 3 hours in the Parker medium at a temperature of 370C. The control group (G1) was incubated in a medium without any hormonal additives,

conditions a decrease of the concentration of prolactin in sheep [34,35].

**of prolactin under** *in vitro* **conditions** 

control of seasonal changes in the sexual activity.


**Table 5.** Effects of the day length and exogenous melatonin on the secretion of prolactin under in vitro conditions.

The concentration of prolactin in the period of lengthening days (March) in the control group was at the level of 145.45 ± 21.2 µg/ml, whereas in the group incubated with melatonin it was lower and amounted to 105.06 ± 11.4 µg/ml. In the period of shortening days (August), the concentration of prolactin in the medium with exogenous melatonin was lower, equaling 45.4 ± 9.2 g/ml, as compared to the control group, being at the level of 65.4 ± 15.7 µg/ml. The lowest concentration of prolactin both in the control group and experimental one was recorded in the short-day period. The experiments conducted confirmed the seasonal rhythm of prolactin, because the highest concentration of this hormone was observed in the period of lengthening days (145.45 ± 21.2 µg/ml). At the same time the lowest one was recorded in the short-day period (13.1 ± 2.8 µg/ml). The experiments conducted under in vitro conditions showed that in the long-day period melatonin exerted a strong inhibitory effect on the secretion of prolactin. The studies confirmed also a direct influence of melatonin on cells of the sheep's pars tuberalis. It was demonstrated that the secretory activity of lactotroph cells of the pituitary gland under in vitro conditions is closely linked to the day length, and at the same time to the secretion of melatonin. It must be stressed at this point that the obtained seasonal distribution of prolactin concentrations in lactating sheep resembles a seasonal rhythm of the prolactin secretion in barren sheep [19]. However, it should be noted that in the group of lactating sheep the secretion of prolactin is much more intensive. Melatonin administered on the 30th day of lactation contributed to the reduction of the prolactin secretion [40].
