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A prolactin (PRL) surge occurs in the female rat during proestrus in response to elevated estradiol levels. The elevated release of ovarian steroids on the day of proestrus is also associated with sexual receptivity. Mating triggers twice-daily PRL surges that supplant the proestrous PRL surge and are responsible for maintaining luteal function during the first half of pregnancy. In order to understand the neuronal mechanisms controlling the proestrous- and mating-induced PRL surges, we examined patterns of Fos expression by immunocytochemistry in specific brain regions as a measure of neuronal activity. Intact female rats were sacrificed at 09.00, 15.00, and 18.00 h on the day of proestrus and the day of diestrus. Brain tissues were also collected at 21.00 h on the day of proestrus from rats receiving intromissions or mounts from males or taken directly from their homecage. On the day of proestrus, the number of neurons expressing Fos in the medial preoptic area (mPOA), medial amygdaloid nucleus (mAMYG), and ventromedial nucleus of the hypothalamus (VMH) was few and was associated with low plasma PRL levels at 09.00 h; however, the number of Fos-positive cells in these brain regions significantly increased at 15.00 and 18.00 h when the proestrous PRL surge occurred. Mating during the evening of proestrus resulted in a dramatic increase in the number of Fos-positive cells in the mPOA, mAMYG, and VMH as well as in the arcuate nucleus (ARC). Nonmated animals in diestrus showed low and unchanged PRL levels and Fos expression in all the brain areas throughout the day. These results suggest that the mPOA, mAMYG, VMH and ARC may be important brain sites for the integration of stimuli associated with proestrous- and mating-induced PRL surges.

Prolactin (PRL) serves an important luteotrophic function in the rat during early pregnancy, expressed as a nocturnal surge in the early morning and a diurnal surge in the late afternoon. Several areas of the hypothalamus, including the preoptic area (POA), the suprachiasmatic nucleus (SCN) and the ventromedial and dorsomedial nuclei (VM-DM) have been implicated in PRL surges. We investigated the temporal relationship between neuronal activity as measured by c-Fos immunocytochemistry in these areas and PRL secretion during early and late pregnancy. Brains were collected at nine time points (24:00, 02:00, 04:00, 06:00, 10:00, 14:00, 16:00, 18:00 and 20:00 h) on days 6–7 and three time points (02:00, 14:00 and 18:00 h) on days 14–15 of pregnancy. Plasma PRL levels determined by radioimmunoassay revealed two surges with peaks at 02:00 and 18:00 h and a trough at 14:00 h on days 6–7, which were absent on days 14–15 of pregnancy. The number of neurons expressing c-Fos in the anterior medial preoptic nucleus, the medial preoptic area and the medial preoptic nucleus, but not the anteroventral preoptic nucleus of the POA, and the VM-DM, showed a semicircadian rhythm which was maximal at 02:00 h or/and 04:00 and 18:00 h and reached the lowest value at 14:00 h, in parallel with the PRL surges in early pregnancy. However, the temporal pattern of c-Fos in these areas was reversed during late pregnancy, with a peak at 14:00 h and low levels at 02:00 and 18:00 h. PRL surges were absent and levels were uniformly low during these times. Neuronal activity in the SCN did not show any correlation with PRL surges. The dorsomedial subdivision of the SCN showed high neuronal activity during the daytime in both stages of pregnancy. Neuronal activity in the ventrolateral subdivision of the SCN was high during the nighttime in early pregnancy, however it exhibited high levels during the daytime in late pregnancy. These results suggest that the two daily surges of PRL secretion during the first half of pregnancy might be related to the temporal rhythm of neuronal activity in the POA and the VM-DM, and a major change in the pattern of neuronal activity in these hypothalamic areas might result in termination of the PRL surges at midpregnancy.

Mating in female rats induces an acute prolactin (PRL) release within 60 min and twice-daily surges of PRL throughout the first 10 days of pregnancy to maintain luteal function. Little is known about the brain mechanism whereby the vaginocervical stimulation is processed to induce PRL release. Our recent results revealed an increase in Fos expression in the arcuate nucleus (ARC) following mating in the intact estrous rat, suggesting that a neuronal network in the brain area may participate in conveying and integrating the genitosensory stimulation. To further investigate the phenotype of activated neurons in the ARC, the present study examined whether beta-endorphin (beta-END) and/or dopamine (DA) neurons are activated by mating, and if so, whether activation is involved in the mating-induced acute release of PRL and the establishment of the twice-daily surges of PRL. In experiment 1, proestrous rats receiving intromissions (mated group) from males or mounts without intromission (mounted group) were sacrificed along with rats taken directly from their home cage (control group) 60 min after the beginning of mating or mounting. Expression of Fos in beta-END neurons and expression of fos-related antigen (FRA) in DA neurons, which were labeled by tyrosine hydroxylase (TH) antibody in the ARC were examined by double-label immunocytochemistry. In experiment 2, proestrous females with indwelling atrial catheters were mated with males. Naloxone (10 microl/min, 2 mg/10 min), an opiate antagonist, or saline was infused before, during and after mating. Blood samples were collected during the mating session and also at several times 3 days after mating. The results showed that mating induced a significant increase in the percentage of beta-END/Fos colabeled neurons and a significant decrease in the number of beta-END cells in all subdivisions of the ARC. In contrast, neither the percentage of FRA/TH colabeled cells nor the number of TH cells was influenced by mating. Mating induced an acute increase in PRL release in saline-treated control animals within 30 min and a subsequent diurnal surge (18.00 h) and a nocturnal surge of PRL (2.00 h) 3 days after mating. Naloxone infusion during mating blocked the mating-induced acute PRL response and the diurnal surge of PRL 3 days after mating, but affected neither the nocturnal surge of PRL nor the incidence of pregnancy. These results demonstrate that (1) beta-END neurons but not DA neurons in the ARC are activated in response to mating in proestrous rats, and (2) the mating-induced activation of beta-END neurons may participate in the acute response of PRL release to mating and the memory mechanism for the establishment of the diurnal PRL surge, but not the nocturnal PRL surge in early pregnancy. These results lead to a conclusion that endogenous opioid peptides may be involved in the neuronal transmission of genitosensory stimulation to induce PRL secretion.