Prolactin release after mating and genitosensory stimulation in females.

Study of the mechanisms by which VCS induces the 10-13 days of twice-daily PRL surges, while at present incompletely understood, will help us to answer several more general neural and endocrine questions: 1) what similarities exist between the suckling-induced and VCS-induced changes in pituitary hormone release, and what central and peripheral mechanisms might be common to both; 2) how do sensory and behavioral factors initiate changes in the pattern of hormone secretion; and 3) what are the mechanisms underlying the establishment of the short-term and long-term mnemonic devices, and do alterations in neural function similar to those responsible for other types of memory underlie this memory? As the data summarized above demonstrate, a neuroethological approach to the study of these questions can be very valuable. The reasons for this are severalfold. First, using naturally occurring behavior as an inducer of PSP, it is possible to use appropriate controls that allow identification of proximal responses that are directly linked to VCS. Females exposed to intromissive stimuli from males show responses that are not shown by females receiving the same flank and perineal somatosensory input from mounts-without-intromission or the olfactory input from nonmating exposure to males. Artificial VCS may induce some nonspecific responses that may be more perceived than real. Second, VCS received by the female during mating is very different from that applied by mechanical or electrical means, and it has been shown that the natural stimuli are important for induction of the PRL surges. The fact that intromissions are normally repeated and intermittent has revealed that the female responds initially with a graded response to these stimuli and that there is a threshold that has to be met for the full response to occur. The set-point of this threshold is influenced by factors that are as yet unknown. Finally, the natural mating condition reveals the contributions of the short-term and long-term mnemonic devices, establishing the existence of a graded to all-or-nothing transition that is required for the occurrence of PSP. In each of these cases, it is clear that these phenomena are obscured when supramaximal artificial stimulation is used as a method to induce PSP. Use of behaviorally appropriate stimulation will continue to be a productive way to study this system.(ABSTRACT TRUNCATED AT 400 WORDS)

Noradrenergic innervation of the ventromedial hypothalamus is involved in mating-induced pseudopregnancy in the female rat.

The ventromedial hypothalamus (VMH) is an oestrogen-responsive area known to facilitate female sexual behaviour in the rat. The VMH is innervated by noradrenergic neurones projecting from the brain stem, and it has been demonstrated that noradrenaline receptor activation in the VMH plays a role in the expression of the lordosis reflex. Noradrenaline has been shown to be released within the VMH after a female receives vaginocervical stimulation (VCS) from the male during mating. VCS also is required to induce twice-daily surges of prolactin (PRL) characteristic of early pregnancy or pseudopregnancy (PSP). To determine whether noradrenaline within the ventrolateral ventromedial hypothalamus (VMHvl) plays a facilitatory role in initiation of PSP, we administered the alpha(1)-noradrenergic receptor agonist, phenylephrine, and the alpha(2)-autoreceptor antagonist, yohimbine, unilaterally into the VMHvl. Phenylephrine stimulated PSP in 85.7% of females given an amount of VCS known to be subthreshold for the induction of PSP, whereas saline infusion (0%) or cannula misplacement (7.7%) were ineffective. Yohimbine had a similar effect, inducing PSP in 85.7% of females, whereas 7.6% of both control groups together showed PSP. Finally, bilateral blockade of alpha(1)-receptors using prazosin blocked PSP in 100% of females given sufficient VCS to induce PSP, whereas saline infusion or misplaced intracerebral cannulae failed to prevent PSP in any animal. In all experiments, vaginal dioestrous was indicative of PSP, in that animals showed a mean number of days between oestrus of 12.8 +/- 0.9. The results of the study demonstrate an important role for the VMHvl in initiation of PSP and suggest that the release of noradrenaline in the VMHvl at the time of mating contributes to neuroendocrine mechanisms responsible for establishing PSP in the female rat.

NMDA-mediated activation of the medial amygdala initiates a downstream neuroendocrine memory responsible for pseudopregnancy in the female rat.

In female rats, genitosensory stimulation received during mating initiates twice-daily prolactin (PRL) surges, a neuroendocrine response that is the hallmark of early pregnancy or pseudopregnancy (P/PSP). Nocturnal and diurnal PRL surges are expressed repeatedly for up to 2 weeks after copulation, suggesting that a neuroendocrine memory for vaginocervical stimulation (VCS) is established at the time of mating. These studies investigated whether the processing and retention of VCS involves acute glutamatergic activation or de novo protein synthesis within the medial nucleus of the amygdala (MEA), a VCS-responsive brain site that is implicated in P/PSP initiation. Pharmacological activation of the MEA with the glutamate agonist, NMDA, initiated nocturnal PRL surges, causing a PSP state in females that had not received VCS. P/PSP initiation by mating was prevented by intra-amygdalar infusion of the NMDA receptor antagonist, 2-amino-5-phosphonopentanoic acid (AP-5), provided that it was administered before mating. AP-5 treatment also disrupted mating-induced c-fos expression in the principle bed nucleus of the stria terminalis and the ventrolateral division of the ventromedial hypothalamic nucleus, but not in the medial or anteroventral periventricular preoptic nuclei. Neither P/PSP nor downstream cellular activation was prevented when a protein synthesis inhibitor, anisomycin, was administered to the MEA. The results indicate that MEA cells are critical to the early processing of VCS through NMDA channel activation, rapidly conveying information to downstream hypothalamic cell groups that modulate neuroendocrine function.

Glutamatergic stimulation of the medial amygdala induces steroid dependent c-fos expression within forebrain nuclei responsive to mating stimulation.

Neurons within the posterodorsal medial amygdala of female rats are known to process vaginocervical stimulation received during mating through N-methyl-D-aspartate channel activation, conveying information to downstream hypothalamic cell groups that modulate neuroendocrine function. Stimulation of these neurons with an excitatory amino acid cocktail of glutamate, aspartate and glycine initiates 10-12 days of prolactin surge secretion that normally are observed only after the receipt of vaginocervical stimulation. Posterodorsal medial amygdala neurons responsive to vaginocervical stimulation also contain estrogen and progesterone receptors. The present experiment examined which downstream sites involved in prolactin secretion show c-fos expression following glutamate receptor activation within the posterodorsal medial amygdala and whether ovarian steroids influence cellular activation in these areas. Ovariectomized female rats implanted with unilateral cannulas directed at the posterodorsal medial amygdala received injections of estradiol benzoate and progesterone or oil before infusion treatment with either excitatory amino acid or control PBS. An additional group of estradiol benzoate+progesterone-treated females was infused with 1.0 microM glycine alone in PBS. Infusions were administered three times at 30 min intervals. FOS induction 90 min after infusion was determined immunohistochemically on the sides ipsilateral and contralateral to the infusion. Of the examined regions, excitatory amino acid treatment and hormone treatment induced three patterns of c-fos expression: 1) responses to both excitatory amino acid and hormone treatment [posterodorsal medial amygdala, medial preoptic area, ventrolateral ventromedial hypothalamic nucleus, bed nucleus of the stria terminalis]; 2) responses to estradiol benzoate+progesterone treatment only [anteroventral periventricular nucleus and dorsomedial nucleus]; and 3) responses to excitatory amino acid only [arcuate nucleus, suprachiasmatic nucleus, and paraventricular nucleus]. These data identify possible circuits by which vaginocervical stimulation, via activation of posterodorsal medial amygdala glutamate-type receptors, initiates and coordinates a series of events within a larger neuroendocrine circuit important for pregnancy.

Effect of neonatal gonadectomy and administration of testosterone on coital masculinization in the ferret.

Male ferrets castrated on postnatal day 5 displayed significantly lower levels of neck gripping, mounting, and pelvic thrusting behavior than groups of males castrated on postnatal days 20 or 35 when tested in adulthood after treatment with testosterone (T). Administering a high dosage of T via sc Silastic capsules to ovariectomized female ferrets over postnatal days 5-20 caused a significant enhancement of all three parameters of masculine coital behavior and ossification of clitorides, in comparison with control females that received no exogenous hormone neonatally. A lower dosage of T given to other ovariectomized females over the same neonatal period caused only slight coital masculinization even though the plasma concentrations of T achieved in females with this particular implant were somewhat higher than the levels normally present in gonadally intact male ferrets at any time between postnatal days 5 and 40. Administration of high and low dosages of T to ovariectomized females over days 20-35 failed to cause any coital or genital masculinization. The results suggest that T acts in the male ferret brain between postnatal days 5 and 20 to cause coital masculinization. The sensitivity of the male brain to T during this period may, however, normally be enhanced by the action of T or some other hormone before day 5.

Plasma concentrations of testosterone and dihydrotestosterone during perinatal development in male and female ferrets.

Concentrations of testosterone (T) and 5 alpha-dihydrotestosterone (DHT) were measured in plasma collected from male and female ferrets at eight perinatal ages, spanning the period when behavioral sexual differentiation occurs in this species. Concentrations of T were significantly higher in males than in females 5 days before birth (day -5) and on postnatal days 10, 15, and 40. Plasma concentrations of DHT were equivalent in both sexes at all ages. In males, mean plasma T (2,278 pg/ml) and DHT (1,989 pg/ml) concentrations were highest on day -5, and declined significantly by postnatal day 5. In females, plasma concentrations of T (1,220 pg/ml) were highest on the day of birth, whereas concentrations of DHT (1,896 pg/ml) were highest on day -5; both declined significantly by postnatal day 5. The mean concentrations of T and DHT in sera from reproductively active adult male ferrets were 26,019 and 888 pg/ml, respectively, whereas sera from seasonally quiescent males contained 2,976 pg/ml T and 252 pg/ml DHT. The results demonstrate that circulating concentrations of T are significantly higher in male than in female ferrets at those neonatal ages when, in other experiments, T administration to females permanently augmented their ability to display masculine coital behavior in adulthood.

Effect of birth on plasma testosterone, brain aromatase activity, and hypothalamic estradiol in male and female ferrets.

The present studies examined the patterns of circulating testosterone (T) within 0-24 h after birth in male and female ferrets along with concomitant changes in neural aromatase activity and hypothalamic concentrations of estradiol (E2). Plasma and brain samples were obtained 0 and 2 h (cesarean delivery) or 0, 2, 12, and 24 h (natural delivery) after birth. Plasma T levels were significantly higher in male neonates 2 h after birth than at 0 h in both cesarean-delivered (9.48 +/- 1.25 vs. 3.37 +/- 0.60 ng/ml) and naturally delivered (19.28 +/- 2.94 vs. 5.13 +/- 1.93 ng/ml) ferrets, while female neonates showed no significant changes in T over these sampling times. T levels had returned to 0 h levels by 12 h in naturally delivered males. T was significantly lower in females than in males 0, 2, and 24 h after natural delivery, whereas T levels were equivalent in males and females immediately after cesarean delivery. Male kits kept on a heating pad for 2 h after natural delivery had lower plasma T levels than males that were left with their mothers over this same period. Brain aromatase activity in anterior hypothalamus-preoptic area, medial basal hypothalamus (MBH), temporal lobe, and cerebral cortex was equivalent in males and females at all postpartum ages, regardless of whether delivery occurred by cesarean section or naturally. However, in naturally delivered kits of both sexes significant elevations in aromatase activity occurred in MBH and temporal lobe 24 h postpartum. Finally, E2 concentrations in anterior hypothalamus-preoptic area and MBH were equivalent 0 and 2 h postpartum in males and females, regardless of whether they were delivered naturally or by cesarean section. The observed postnatal elevation in T may contribute to brain and behavioral sexual differentiation of male ferrets. It is unclear, however, whether such an effect of T depends on its neural aromatization to E2.

Sex difference in the effect of mating on the pulsatile secretion of luteinizing hormone in a reflex ovulator, the ferret.

Sex differences in the pulsatile secretion of LH were examined in male and female ferrets after mating. Female ferrets which were either gonadally intact and in estrus or gonadectomized and maintained on a pulsed regimen of daily estradiol (E2) injections exhibited a prolonged rise in plasma LH, characterized by an elevation in mean LH levels and an increase in the number of LH pulses after receiving an intromission from a stud male. By contrast, no such increase in LH secretion occurred in males which achieved an intromission with a female, regardless of whether they were gonadally intact and in breeding condition or gonadectomized and given pulsed estrogen. In fact, intact breeding males which achieved an intromission had significantly fewer LH pulses 1-5 h later than unmated males bled serially over the same time period. This decrease in LH pulse frequency was followed by a significant rise in mean plasma levels of androgen 5-12 h later. When a sexually dimorphic LH response to intromission was observed in gonadectomized E2-treated ferrets, we asked whether this could reflect a sex difference in pituitary responsiveness to the endogenous release of GnRH. Thus, plasma LH levels were measured in gonadectomized and gonadectomized E2-treated ferrets for 2 h after iv injection of GnRH. In the absence of gonadal steroids, ferrets exhibited a sex-specific difference in LH responsiveness to GnRH; however, no sex difference was apparent under the influence of E2. These findings demonstrate that ferrets' sexually dimorphic LH responses to intromission probably reflect a sex difference in the processing of somatosensory inputs from the genitalia or in the neural control of GnRH release into the pituitary portal vessels.

Diurnal fluctuations in mating-induced oxytocinergic activity within the paraventricular and supraoptic nuclei do not influence prolactin secretion.

Previous studies have implicated oxytocin (OT) in the control of surge-type PRL secretion in the pregnant and pseudopregnant rat. The present studies examined the relationship between mating-induced activation of OT neurons in the paraventricular (PVN), supraoptic (SON), and anterior commissural (ACN) nuclei and PRL secretion. Activity within OTergic neurons, as measured by increased c-fos expression, was examined immediately and 5 days following mating in ovariectomized, estrogen-plus-progesterone-treated rats at the time when nocturnal PRL surges are expressed (0600 h) and at an intersurge time (2400 h). Females received fifteen intromissions (15I), 15 mounts-without-intromission (MO), or no stimulation (homecage, HC) from a sexually experienced male. Receipt of 15I at 0600 h induced significantly higher numbers of OT immunoreactive (OT-IR) cells and FOS/OT-IR double-labeled cells in the parvocellular division of the PVN (PVNparv) and in the SON than did 15I at 2400 h. Numbers of OT-IR and FOS/OT-IR cells in the ACN and in the magnocellular compartment of the PVN (PVNmag) were not influenced by mating at either time. In contrast, acute PRL secretion induced within 5-30 min by 15I was not influenced by whether mating occurred at 1800 h (diurnal surge), 2400 h, or 0600 h, nor were plasma OT levels elevated during the 1 h following 15I or MO at these times. Examination of FOS-IR cells throughout the hypothalamus across the two times of day revealed previously unreported differences between 15I and control MO treatments in the PVN, SON, and the ventrolateral part of the arcuate nucleus (ARCvl). On day 5 post mating, numbers of OT-IR and FOS/OT-IR cells in the PVN, SON, and ACN were very low and were similar between 0600 h and 2400 h and between females that showed (15I) or did not show (MO) mating-induced PRL surges characteristic of pregnancy. The results of these studies demonstrate that intromissive but not mounts-only stimulation from males induces a rapid increase in OT-IR staining and OT neuron activation in the PVNparv and the SON. These mating-induced responses in OT neurons occurred within 1 h after mating only at 0600 h, suggesting a diurnal fluctuation in sensitivity to intromissive stimulation. Changes in OTergic function were not seen in response to mating at other times of day, nor at the time of the nocturnal PRL surge 5 days after mating. We conclude that OT activity induced by mating does not act to stimulate PRL secretion directly, but may be involved in the process(es) by which genitosensory stimulation initiates surge-type PRL secretion.

Pseudorabies virus tracing of neural pathways between the uterine cervix and CNS: effects of survival time, estrogen treatment, rhizotomy, and pelvic nerve transection.

The transneuronal tracer, pseudorabies virus (PRV), was used to identify pathways from the uterine cervix which may be involved in induction of analgesia and abbreviation of estrus by vaginocervical stimulation. In Experiment I, PRV immunoreactivity (PRV-IR) in brain and spinal cord was examined 3-5 days after injection into the cervix of ovariectomized (OVX) female rats given estrogen (E) or control treatments. No differences in viral labeling were observed between OVX and OVX+E females at any time. PRV-infected cells were observed to increase as a function of time and at progressively higher CNS levels. PRV-IR neurons were first observed on day 3 post-infection at L6 in the SPN. Increased labeling was observed at day 4 in the SPN and the DGC at L6 and S1 spinal segments. Dorsal horn neurons showed PRV-IR by 4.5 days. Five days post-infection, labeling was seen in the IML and lamina X in T12-L1 segments, and in medullary raphe, A5, nPGi, nGi, DMV, lateral reticular, Barrington's nuclei, and in the midbrain PAG. In Experiment II, the effects of bilateral L6 dorsal root rhizotomy (RH) combined with unilateral (UPx) or bilateral (BPx) pelvic nerve transection on PRV infectivity were examined 5 days after infection. Despite reductions in substance P labeling in the dorsal horn following RH, PRV-IR neurons persisted in this area. In RH+UPx females, labeling persisted bilaterally in the SPN and DGC at L6. RH+BPx almost completely eliminated the PRV labeling in L6 and S1. Horizontal sections showed distinct patterns of infectivity within the IML of thoracolumbar and SPN of lumbosacral segments consistent with infection in the hypogastric and pelvic nerves, respectively. Our data indicate that retrograde transport of PRV occurs via the hypogastric and pelvic nerves after injection of the virus into the uterine cervix. Furthermore, significant intraspinal processing is likely to occur between thoracolumbar and lumbosacral levels in the modulation of reproductive tract function.

Vaginocervical stimulation suppresses the expression of c-fos induced by mating in thoracic, lumbar and sacral segments of the female rat.

In female rats, vaginocervical stimulation induces neuroendocrine responses necessary for pregnancy as well as analgesia to a variety of noxious stimuli. In this study, Fos immunocytochemistry was used to detect vaginocervical stimulation-induced changes in the activity of spinal neurons at levels T11-S3, segments known to receive afferent input from nerves which innervate the reproductive tract. Adult ovariectomized estrogen and progesterone-treated rats were killed 1 h after receiving mating stimulation from males, which included five or 15 intromissions, mounts-without-intromission by use of either vaginal masks or genitally-anaesthetized males, or immediately after being removed from their home cages. At all spinal levels, Fos labelling was lowest in the home cage group (50 +/- 22 cells), intermediate in the groups receiving intromissions (84 +/- 8 and 118 +/- 22 cells) and highest in groups receiving mounts-without-intromission stimulation (187 +/- 21 and 218 +/- 35 cells). Significant increases above control levels following intromissive stimulation were observed at levels L6, S1 and S2. Surprisingly, both groups receiving mounts-without-intromission showed significantly higher numbers of Fos-positive cells than did the fully mated groups at all levels. Analysis of selected spinal segments by Rexed's laminae revealed that intromissive stimulation increased Fos labelling above control levels in laminae II-V and X at L6, and laminae I, II, V and X at S1; vaginocervical stimulation did not increase labelling at L1. The greater Fos responses seen in mounts-without-intromission animals than in control or intromitted animals were apparent at L1, L6 and S1 within the same laminae (II-V and X). These results suggest that stimulation of the uterine cervix initiates activity within L6-S2 neurons which receive pelvic nerve afferents and that such stimulation suppresses activity at all levels within populations of neurons normally activated by cutaneous somatic inputs received from male mounts. As antinociceptive agents are known to suppress c-fos expression, vaginocervical stimulation received during natural mating may be capable of initiating spinal and/or brain mechanisms of analgesia.

Plasma concentrations of oestradiol and oestrone during perinatal development in male and female ferrets.

The concentrations of oestradiol and oestrone in peripheral plasma of male and female ferrets 5 days before and 7, 15 and 30 days after birth were measured. Both steroids were present in high concentrations prenatally. Much lower levels were found in samples collected on day 7 and later, when the concentrations were similar to those of adult gonadectomized animals. No significant sex difference was seen for the concentration of either steroid at any age studied. These results, and those previously reported showing the absence of a circulating binding protein and the presence of oestradiol receptors in the hypothalamus in the perinatal period in this species, suggest that brains of both males and females are exposed to significant amounts of oestrogen during development. These findings lend support to the possibility that prenatal exposure to oestrogen plays a role in organizing the potential for female behaviour in male and female ferrets.

Metabolism of dihydrotestosterone to 3 alpha-androstanediol in brain and plasma: effect on behavioural activity in female rats.

Six experiments were carried out to determine whether dihydrotestosterone (5 alpha-androstan-17 beta-ol-3-one; DHT) acts to inhibit oestradiol (OE2)-induced lordosis behaviour after its metabolic conversion to 5 alpha-androstane-3 alpha,17 beta-diol (3 alpha-androstanediol, 3 alpha-Adiol). In experiments 1 and 2, ovariectomized rats were treated with several doses of DHT or 3 alpha-Adiol, injected with OE2 and progesterone, and tested for lordosis responsiveness. Significant inhibition of lordosis occurred after a dose of 3 alpha-Adiol which was approximately threefold less than the effective DHT dose. In experiments 3 and 4, plasma concentrations of DHT and 3 alpha-Adiol were measured after the injection of these steroids to ovariectomized rats at doses shown to be both sufficient or insufficient to inhibit lordosis. Behaviourally effective dosages of DHT and 3 alpha-Adiol produced circulating concentrations of 3 alpha-Adiol greater than those produced by behaviourally ineffective doses of DHT or 3 alpha-Adiol. At 30 min after injection of DHT (experiment 3), 78.8% of plasma androgens were in the form of 3 alpha-Adiol, while after injection of 3 alpha-Adiol, only 7.4% were DHT. When plasma DHT and 3 alpha-Adiol were measured at 3, 6, 9 and 12 h after steroid injection (experiment 4), plasma levels of 3 alpha-Adiol produced by the behaviourally subthreshold dose of DHT were significantly lower than levels produced by behaviourally sufficient dosages of DHT or 3 alpha-Adiol. In experiments 5 and 6, concentrations of DHT and 3 alpha-Adiol were measured in five brain regions 1 and 6 h after injection of behaviourally sufficient doses of these steroids to ovariectomized females. At 1 h after injection, similar levels of DHT and 3 alpha-Adiol were measured in DHT- and 3 alpha-Adiol-injected females, and DHT concentrations in the preoptic area were significantly higher in both groups than in any other brain area. At 6 h, animals injected with DHT had significantly higher levels of DHT in all brain areas combined than did 3 alpha-Adiol- or vehicle-injected animals. Brain concentrations of 3 alpha-Adiol were not different between groups injected with DHT, 3 alpha-Adiol or vehicle at this time. In brain, 34.6% of DHT had been converted to 3 alpha-Adiol after 1 h and 53.0% of 3 alpha-Adiol had been converted to DHT.(ABSTRACT TRUNCATED AT 400 WORDS)

Failure of the synthetic androgen 17 beta-hydroxy-17 alpha-methyl-estra-4,9,11-triene-3-one (methyltrienolone, R1881) to duplicate the activational effect of testosterone on mating in castrated male rats.

Administration of the synthetic steroid, 17 beta-hydroxy-17 alpha-methyl-estra-4,9,11-triene-3-one (methyltrienolone, R1881), which is an androgen receptor agonist not metabolized in vivo, at doses of 400, 800 or 2400 micrograms/kg s.c. in propylene glycol stimulated mounting and ejaculation only slightly in sexually experienced castrated rats. A similar low level of mating was observed in a group of castrated rats given testosterone at doses of 400 followed by 800 micrograms/kg. However, when treated with a higher dose of testosterone (2400 micrograms/kg) these castrated rats displayed significantly higher levels of mounting and ejaculation than rats treated with any of the doses of R1881. Also, when this higher dosage of testosterone was substituted for each dose of R1881, significant increments in mounting and ejaculation occurred in all groups. These findings show that R1881 is only marginally effective in restoring sexual behaviour in castrated rats, suggesting that the activation of neural androgen receptors cannot by itself account for the activational effect of testosterone on mating behaviour in gonadally intact male rats.

Absence of a diurnal rhythm in lordosis behaviour induced by oestrogen in gonadectomized rats.

Gonadectomized rats bearing s.c. Silastic capsules containing crystalline oestradiol-17 beta diluted with cholesterol, or oestradiol-17 beta dissolved in sesame oil were tested for the presence of a diurnal rhythm in the display of lordotic behaviour. In experiment 1, female rats received four consecutive tests at intervals of 8 h in a lighting regimen of 12 h light: 12 h darkness beginning 4, 14 and 28 days after implantation of 5 mm capsules of oestradiol. After a single test on day 4, male rats were tested on days 14-15 only, at the same times as the female rats. Female animals were tested while vaginal-cervical stimulation was prevented by vaginal masking beginning 35 days after implantation of oestradiol. In experiment 2, lordotic responsiveness of female rats was assessed beginning 4 days after implantation of oestradiol once on each of 3 consecutive days, with each test occurring at a different time of day. Finally, in experiment 3, female rats were tested as in experiment 1 beginning 4 dys after implantation of lower threshold amounts of oestradiol in oil-filled capsules. In no experiment were changes in lordotic behaviour observed as a function of the time of day. These findings failed to support recent reports of a sexually dimorphic rhythm in lordotic responsiveness to oestradiol in the rat.

Effects of differential mating stimulation on the onset of prolactin surges in pseudopregnant rats.

In the female rat, stimulation of the uterine cervix (CS) during mating or by artificial means induces daily diurnal and nocturnal surges in prolactin (PRL) secretion which, in the absence of fertilization, result in an 12-day anestrous period called pseudopregnancy (PSP). The amount or type of mating stimulation received by the female during mating determines whether or not PSP occurs, but it has not yet been determined whether different amounts of mating stimulation can alter the time of onset of PRL surges. The present studies examined the latency in days to the first nocturnal PRL surge following mating. Plasma PRL was measured in samples obtained via intra-atrial catheters at 0200-0500h on the day of estrus (Day 0) and on the subsequent 3 days (Days 1-3). In Experiment 1, proestrus females received mating stimulation which was more than (15 intromissions, 15I) or less than (mounts-without-intromission only, MO) sufficient to induce PSP. Surges were absent in 15I females until Days 1-2 and in MO females on all days. In Experiment 2, females received five intromissions (5I) in paced and nonpaced mating tests, types of mating treatments which were expected to induce PSP in some but not all females. PRL surges were not evident at any sampling time in females that continued to cycle, while PRL surges occurred consistently on Day 2 among PSP females. Among PSP females, those receiving 5I showed significantly higher PRL on Day 0 than did 15I females. In both experiments, plasma progesterone concentrations were not higher in PSP than in non-PSP animals until Day 3. In Experiment 3, PRL levels in single samples obtained by cardiac puncture on Day 0 were similar to those seen in the first two experiments. Thus, PRL secretion at the time of the first postmating nocturnal surge is influenced by the type of CS received some 8-10 h earlier. However, if sufficient CS is received to induce the neural changes of PSP, the nocturnal PRL surges are expressed in an all-or-none fashion by 1-2 days after mating.

Effects of paced and non-paced mating stimulation on plasma progesterone, 3 alpha-diol and corticosterone.

In the female rat, gonadal and adrenal progestins and androgens modulate sexual receptivity and in turn, their levels increase in response to mating stimulation. Paced mating, in which the female controls the timing of sexual contacts with the male, is particularly effective at eliciting acute increases in progesterone (P) and 5 alpha-Androstane-3 alpha, 17 beta-diol (3 alpha-Diol). Interestingly, restraint stress produces comparable increases in P and 3 alpha-Diol levels, as well as increases in corticosterone (CORT) levels. In this study, we explored the possibility that paced mating would be associated with increased CORT, in conjunction with mating-induced increases in P and 3 alpha-Diol. Ovariectomized rats primed with estradiol benzoate (10 micrograms in oil SC) and P (0.5 mg in oil s.c.) received a single ejaculatory series from males in paced or non-paced mating tests. Fifteen minutes post-mating rats were exposed to CO2 and rapidly decapitated for the collection of trunk blood and determination of P, 3 alpha-Diol and CORT via radioimmunoassay. As expected, P and 3 alpha-Diol concentrations were significantly elevated in serum obtained from animals allowed to pace their sexual contacts with males compared to those which did not pace their contacts. Importantly, although all mated animals had CORT levels between 10-20 micrograms/dl, there were no differences between paced and non-paced conditions. This suggests that the acute rises in P and 3 alpha-Diol in response to paced mating are not due to paced mating being more stressful than non-paced mating.

Excitotoxic lesions of the medial amygdala differentially disrupt prolactin secretory responses in cycling and mated female rats.

This study examined the role of the posterodorsal medial amygdala (MePD) in the control of prolactin secretion in gonadally intact female rats 20 min after mating, during the oestrous cycle, and during early pregnancy/pseudopregnancy (P/PSP). Cycling females received bilateral infusions of an excitotoxic dose of N-methyl-D-aspartate (NMDA) or vehicle into the MePD. Two to 4 weeks later, they were surgically implanted with intra-atrial catheters for repeated blood sampling and were mated on the evening of proestrus until receiving 5, 10, 15 or 20 intromissions or 15 mounts-without-intromission (MO) from males. The percentages of rats becoming P/PSP increased as a function of numbers of intromissions received. All groups receiving intromissions showed similar approximately four-fold increases in plasma prolactin concentrations 20 min after mating, while MO rats showed no increase at this time. There was no effect of NMDA lesion on this acute secretory response. Among rats that continued cycling, bilateral MePD lesion completely abolished the diurnal preovulatory prolactin surge, while incomplete and sham lesions did not. In rats that subsequently became P/PSP, bilateral lesion of the MePD resulted in dampening of prolactin concentrations at all sampling times 6-7 days after mating, while incomplete or sham lesions did not alter prolactin secretion at these times. These results demonstrate that the MePD is selectively important for the secretion of prolactin on the afternoon of proestrus and that this structure may also modulate prolactin release in P/PSP rats. Activity within MePD neurones does not appear to be required for the acute prolactin response to vaginocervical stimulation which occurs within minutes after mating.

Acute luteinizing hormone and prolactin responses to paced mating stimulation in the estrous female rat.

The present experiments sought to characterize the particular stimuli received during mating in the female rat which induce acute increases in luteinizing hormone (LH) and prolactin (PRL) following copulation. Comparisons were made between cycling females mated on the evening of proestrus in partitioned chambers in which spontaneous patterns of approach/withdrawal toward the male served to pace copulatory stimulation (paced), in non-partitioned chambers in which female regulation of intervals between copulatory mounts was prevented (non-paced), or under conditions in which they received mounts-without-intromission (mounts-only). Frequent blood samples were withdrawn via surgically implanted intra-atrial catheters. In experiment 1, blood samples for LH determinations were taken at 15-min intervals for 1 h prior to and for 2 h after mating on the evening of proestrus. In experiment 2, samples for PRL determinations were taken at 10-min intervals for 30 min prior to and for 90 min after mating on proestrus and at 0300, 0400 and 0500 h on the day of estrus (reported times corrected for reversed light cycle). LH levels were significantly higher in paced animals 15 min after initiation of mating than in non-paced and mounts-only females; no differences in LH were seen between females who subsequently became or did not become pregnant/pseudopregnant (P/PSP). PRL values were not different between groups receiving the different mating treatments at any time; however, P/PSP animals showed significantly higher levels of PRL between 20 to 60 min after mating than did non-P/PSP females. No differences in PRL were seen between mating treatments or pseudopregnancy condition at 0300 to 0500 h on estrus. Paced females in both experiments received intromissions at a significantly slower rate than did non-paced females. There was a significant positive correlation (r = 0.619, P<0.001) between LH concentration at 15 min and the inter-intromission interval (in seconds) in paced and non-paced groups of females. These data suggest that an LH response to mating is dependent upon the particular characteristics of mating stimulation received. In addition, they demonstrate that PRL increases acutely after mating stimulation in animals destined to become P/PSP but does not increase in response to those characteristics of mating stimulation which induced increases in LH.

Effects of paced mating on c-fos gene expression in the female rat brain.

When estrous female rats control or pace (P) their sexual contacts with males, several neuroendocrine and behavioral responses to mating occur that are not observed or are greatly attenuated after nonpaced mating. The present study examined whether the distribution and amount of FOS immunoreactivity (FOS-IR) induced in brain by mating would be altered in females receiving paced rather than nonpaced mating stimulation. In the first experiment, females received 5 or 15 intromissions during paced mating tests (5P and 15P), 5 or 15 intromissions during nonpaced mating tests (5NP and 15NP), 15 mounts-without-intromission (MO) or remained in their homecages (HC). Selective increases 1 h after paced mating stimulation were observed in the posterodorsal medial amygdala (MePD), where significantly more FOS-IR cells were present in the 5P and 15P groups than in the respective NP groups. The 5P, 5NP and 15NP had significantly more FOS-IR than the HC, MO, and 5NP groups, and the 5P group had levels of FOS-IR which were equivalent to that seen in the 15NP group. In the posteromedial portion of the bed nucleus of the stria terminalis (BNSTpm) and the ventrolateral portion of the ventromedial nucleus of the hypothalamus (VMHvl), paced mating induced significantly greater numbers of FOS-IR cells than did either MO or HC treatments; increases induced by nonpaced mating were not statistically greater than HC controls. No differences between groups were seen in the medial preoptic area (mPOA). In the second experiment, experimentally lengthening the interintromission interval (III) as well as increasing the intromission duration to mimic the characteristics of paced mating, resulted in significant increases in FOS-IR in the MePD but not in the other three brain regions. These results demonstrate that paced mating is more effective in inducing c-fos expression than nonpaced mating, and that the MePD is particularly sensitive to differing characteristics of the mating stimuli received.