Insulin but not leptin protects olfactory mucosa from apoptosis.

The mammalian olfactory mucosa (OM) is continually renewed throughout life. Owing to their position in the nasal cavity, OM cells are exposed to multiple insults, including high levels of odourants that can induce their death. OM regeneration is therefore essential to maintain olfactory function, and requires the tight control of both cell death and proliferation. Apoptosis has been implicated in OM cell death. Olfaction is one of the senses involved in food intake and depends on individual nutritional status. We have previously reported the influence of hormones related to nutritional status on odour perception and have shown that the OM is a target of insulin and leptin, two hormones known for their anti-apoptotic properties. In the present study, we investigated the potential anti-apoptotic effect of these metabolic hormones on OM cells. Both Odora cells (an olfactive cell line) and OM cells treated with etoposide, a p53 activity inducer, exhibited mitochondrial-dependent apoptosis that was inhibited by the pan-caspase inhibitor zVAD-fmk. Insulin, but not leptin, impaired this apoptotic effect. Insulin addition to the culture medium reduced p53 phosphorylation, caspase-3 and caspase-9 cleavage, and caspase-3 enzymatic activity induced by etoposide. The apoptotic wave observed in the OM after interruption of the neuronal connections between the OM and the olfactory bulb by bulbectomy was impaired by intranasal insulin treatment. These findings suggest that insulin may be involved in OM cellular dynamics, through endocrine and/or paracrine-autocrine effects of circulating or local insulin, respectively.

Endothelin evokes distinct calcium transients in neuronal and non-neuronal cells of rat olfactory mucosa primary cultures.

The olfactory system is regulated by several nervous and hormonal factors, and there is a growing body of evidence that some of these modulations already take place in the olfactory mucosa (OM). We recently suggested that, among others, vasoactive peptides might play multifaceted roles in different OM cells. Here we studied the effect of the vasoconstrictive peptide endothelin (ET) in the rat OM. We identified different components of the ET system both in the olfactory mucosa and in long-term primary culture of OM cells, composed of olfactory sensory neurons (OSNs) lying on a blend of non-neuronal OM cells (nNCs). We demonstrated that ET receptors are differentially expressed on OM cells, and that ET might be locally matured by the endothelin-converting enzyme ECE-1 located in OSNs. Using calcium imaging, we showed that ET triggers robust dose-dependent Ca(2+) responses in most OM cells, which consist of a transient phase, followed, in nNCs, by a sustained plateau phase. All transient responses depended on intracellular calcium release, while the sustained plateau phase also depended on subsequent external calcium entry. Using both pharmacology and spotting lethal (sl/sl) mutant rats, lacking functional ET(B) receptors, we finally demonstrated that these effects of ET are mediated through ET(B) receptors in OSNs and ET(A) receptors in nNCs.The present study therefore identifies endothelin as a potent endogenous modulator of the olfactory mucosa; specific endothelin-mediated Ca(2+) signals may serve distinct signaling functions, and thereby suggest differential functional roles of endothelin in both neuronal and non-neuronal OM cells.

Nutritional status modulates behavioural and olfactory bulb Fos responses to isoamyl acetate or food odour in rats: roles of orexins and leptin.

Food odours are major determinants for food choice, and their detection depends on nutritional status. The effects of different odour stimuli on both behavioural responses (locomotor activity and sniffing) and Fos induction in olfactory bulbs (OB) were studied in satiated or 48-h fasted rats. We focused on two odour stimuli: isoamyl acetate (ISO), as a neutral stimulus either unknown or familiar, and food pellet odour, that were presented to quiet rats during the light phase of the day. We found significant effects of nutritional status and odour stimulus on both behavioural and OB responses. The locomotor activity induced by odour stimuli was always more marked in fasted than in satiated rats, and food odour induced increased sniffing activity only in fasted rats. Fos expression was quantified in periglomerular, mitral and granular OB cell layers. As a new odour, ISO induced a significant increase in Fos expression in all OB layers, similar in fasted and satiated rats. Significant OB responses to familiar odours were only observed in fasted rats. Among the numerous peptides shown to vary after 48 h of fasting, we focused on orexins (for which immunoreactive fibres are present in the OB) and leptin, as a peripheral hormone linked to adiposity, and tested their effects of food odour. The administration of orexin A in satiated animals partially mimicked fasting, since food odour increased OB Fos responses, but did not induce sniffing. The treatment of fasted animals with either an orexin receptors antagonist (ACT-078573) or leptin significantly decreased both locomotor activity, time spent sniffing food odour and OB Fos induction in all cell layers, thus mimicking a satiated status. We conclude that orexins and leptin are some of the factors that can modify behavioural and OB Fos responses to a familiar food odour.

Expression of insulin system in the olfactory epithelium: first approaches to its role and regulation.

Food odours are major determinants for food choice; their detection is influenced by nutritional status. Among different metabolic signals, insulin plays a major role in food intake regulation. The aim of the present study was to investigate a potential role of insulin in the olfactory mucosa (OM), using ex vivo tissues and in vitro primary cultures. We first established the expression of insulin receptor (IR) in rat olfactory mucosa. Transcripts of IR-A and IR-B isoforms, as well as IRS-1 and IRS-2, were detected in OM extracts. Using immunocytochemistry, IR protein was located in olfactory receptor neurones, sustentacular and basal cells and in endothelium of the lamina propria vessels. Moreover, the insulin binding capacity of OM was quite high compared to that of olfactory bulb or liver. Besides the main pancreatic insulin source, we demonstrated insulin synthesis at a low level in the OM. Interestingly 48 h of fasting, leading to a decreased plasmatic insulin, increased the number of IR in the OM. Local insulin concentration was also enhanced. These data suggest a control of OM insulin system by nutritional status. Finally, an application of insulin on OM, aiming to mimic postprandial insulin increase, reversibly decreased the amplitude of electro-olfactogramme responses to odorants by approximately 30%. These data provide the first evidence that insulin modulates the most peripheral step of odour detection at the olfactory mucosa level.

Transcription profile analysis reveals that OBP-1F mRNA is downregulated in the olfactory mucosa following food deprivation.

Neuroanatomical data show that olfactory mucosa (OM) is a possible place for interactions between nutrition and smell. A combination of differential display mRNA analysis together with a macroarray screening was developed to identify transcripts that are differentially expressed in rat OM following food deprivation. Using this method, backed on a stringent statistical analysis, we identified molecules that fell into several Gene Ontology terms including cellular and physiological process, signal transduction, and binding. Among the 15 most differentially expressed molecules, only one was upregulated, but 14 were downregulated in the fasted state among which was, unexpectedly, odorant-binding protein 1F (OBP-1F). Because of its potential relevance to olfactory physiology, we focused our further analysis on OBP-1F using in situ hybridization, quantitative polymerase chain reaction, and western blot analysis. OBP-1F was highlighted in the lateral nasal glands, but its expression (mRNA and protein) did not change following food deprivation. Only the minor fraction of OBP-1F mRNA expressed by the OM itself was downregulated following 48 h fasting. Altogether, our results suggest that the fine transcriptional control of OBP-1F in the OM following food deprivation could be efficient only at the local level, close to its site of secretion to participate in the perireceptor events of the olfactory signal reception.

Endothelial and neuronal nitric oxide synthases are present in the suprachiasmatic nuclei of Syrian hamsters and rats.

Nitric oxide (NO) is involved in the transmission of light information to suprachiasmatic nuclei (SCN). By immunocytochemistry, we showed that both neuronal and endothelial NO synthase isoforms (nNOS and eNOS) were present in the SCN of rats and hamsters. nNOS-immunoreactive neurons were located mainly around the SCN with only a few nNOS neurons within the nucleus. By double-label immunocytochemistry, we also found, within the population of SCN glial fibrillary acidic protein (GFAP)-immunoreactive astrocytes, a subpopulation of eNOS-immunoreactive astrocytes. Using Western blot analysis, we detected in SCN protein extracts eNOS and nNOS proteins having the expected 140 and 150 kDa molecular weights, respectively. By in situ hybridization of a 2.4-kb murine eNOS probe, mRNA for eNOS was located in the SCN of rats and hamsters. The transcript was further identified by detection of a RT-PCR product of the predicted size, after amplification of total RNA with primers specific for eNOS. In the SCN and cerebellum, the size of the mRNA for nNOS, detected with a rat probe on Northern blot, was approximately 10.5 kb, corresponding to that previously published. In the same tissues, we found two transcripts, one weakly expressed at approximately 4.0 kb and another more strongly expressed at approximately 2.6 kb, both hybridizing with two non-overlapping murine and rat eNOS probes. These results suggested the existence in the SCN of alternate transcripts for eNOS. We propose that two pathways could link light stimuli and NO release in the SCN: one involving N-methyl-D-aspartate (NMDA) receptors and nNOS in neurons; the other linking alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptors and eNOS in astrocytes.

Long-term exposure of hypothalamic explants to melatonin alters the release of gonadotrophin releasing hormone and the density of melatonin binding sites in the pars tuberalis of the male mink (Mustela vison).

To investigate the action of melatonin on the reproductive system, the effect of prolonged versus short-term exposure to melatonin on the release of gonadotrophin releasing hormone (GnRH) was examined in hypothalamic explants of male mink sacrificed in July, September or November. Mediobasal hypothalamic (MBH) explants including the pars tuberalis (PT) were incubated for 1 night with or without melatonin (10(-8) M) for 8 hr or 16 hr and the release of GnRH was then measured. The next day, the explants were incubated further but in a melatonin free buffer, and the release of GnRH was measured with increasing time. Half of the July and September explants had melatonin binding sites quantified by autoradiography. In November, a 16-hr exposure to melatonin induced a significant increase in the release of GnRH during the night, compared with control or 8-hr melatonin exposure. This increase persisted for at least 45 min after the withdrawal of melatonin, suggesting a stimulatory effect of melatonin on the synthesis of GnRH; this effect was apparent in July, September and November. In September, the density of melatonin binding in the PT was significantly lower in the explants incubated for 16 hr with melatonin, compared with those incubated for 8 hr. Thus, in vitro, a long exposure to melatonin, mimicking a single long night, stimulates the release and synthesis of GnRH in parallel with a decrease in the density of melatonin binding in the PT. These effects seem to depend heavily on the duration of exposure to melatonin.

Effect of short photoperiods on the in vitro GnRH release by hypothalamic explants in intact and castrated male Syrian hamsters: relation to testicular regression and recrudescence.

Prolonged exposure of adult Syrian hamsters to short days decreases LH and FSH circulating levels within 2-4 weeks, then induces testicular regression. After 18 weeks of short days, the testis size and gonadotropin levels increase spontaneously. This study investigated whether these phases of photosensitivity and photorefractoriness corresponded to variations of in vitro GnRH release. Male hamsters were either kept under long days (LD 16:8) or transferred to short days (SD 6:18) and sacrificed from 2-26 weeks after transfer. To separate the effects of testis feedback from a possible direct photoperiodic drive on the hypothalamus, males were bilaterally castrated, kept under LD or transferred to SD, and sacrificed from 2-14 weeks after transfer. Hypothalamic explants were incubated in a saline buffer for three periods of 15 min and exposed to KCl (60 mM) for 15 min. The return to basal values was followed for six periods of 15 min, then the explants were stimulated with copper complexed equimolarly with histidine (Cu/His, 200 microM) and prostaglandin E2 (PGE2, 10 microM). At the end of the incubation period, the concentration of GnRH remaining in the explants was measured. In intact males, GnRH release in vitro increased significantly between 2 and 4 weeks after transfer to short days; it returned to values similar to LD ones between 6 and 12 weeks, during the phase of testis involution. At the beginning of photorefractoriness (SD 14-18), it increased transiently and returned to values similar to LD ones from SD 20, during the testis spontaneous recrudescence. After castration, the in vitro GnRH release decreased significantly under LD and SD. The transfer of castrated hamsters to SD resulted in transient increases of GnRH release (SD 4, 8 and 14), and in a progressive loss of the explant's ability to release GnRH in vitro. These results showed a photoperiodic regulation of in vitro GnRH release and a testis feedback effect on this release. They demonstrated an inverse relationship between the readily releasable pool of GnRH and the circulating levels of gonadotrophins at the beginning of photosensitive and photorefractory phases and after castration.

Seasonal variation of melatonin binding sites in the pars tuberalis of the male mink (Mustela vison).

The seasonal changes of 2-[125I]iodomelatonin binding were studied using quantitative autoradiography in the pars tuberalis (PT) of the mink, a short-day breeder, kept out of doors. Studies were performed at 7 times of the year (July, September, October, January, February, and May), corresponding to different states of responsiveness of the gonadal system to the photoperiod. Melatonin binding was observed in the PT and on the ventral border of the pars distalis. Histological staining revealed that the binding on the border of the pars distalis corresponded to the zona tuberalis, a ventral extension of the PT. The binding was specific and saturable. The density of melatonin binding varied significantly with the time of year. The lowest density of binding was found in July, when animals experienced a long daylength and sexual rest, increased from July to reach a maximum in October, when animals experienced decreasing daylength and the hypothalamo-pituitary activity resumed, then slightly decreased and remained constant from November to May. The saturation study demonstrated that the decrease in melatonin binding density between October and February resulted from a change in the number (Bmax: October 70.6 +/- 4.0 vs February 49.6 +/- 2.8 fmol/mg protein; P < 0.01) but not in the affinity (Kd: October 33.6 +/- 7.1 vs February 20.8 +/- 5.1 pM; P > 0.05) of the binding sites. These results are discussed according to the different phases of mink reproductive cycle and to reported data on the sites of action of melatonin on seasonal reproduction and prolactin secretion.

Effect of melatonin on the release of gonadotropin-releasing hormone and cyclic AMP from the rat hypothalamus: an in vitro study.

Using an in vitro static incubation system of adult male rat hypothalami, we have studied the effect of melatonin on the release of gonadotropin-releasing hormone (GnRH) and cyclic adenosine monophosphate (cAMP). Mediobasal hypothalamus (MBH) and preoptic area (POA) were incubated separately in Minimum Essential Medium (MEM) for 6 h. The release of GnRH was measured by radioimmuno-assay in the incubation medium sampled every 7.5 min. In the MBH and POA incubation medium, the mean amount of GnRH released was 8.9 +/- 1.1 and 3.4 +/- 0.6 pg GnRH/7.5 min, respectively (P < 0.01). The mean number of GnRH pulses under basal conditions was 2 +/- 0.3 per 2 h in the MBH and 1.6 +/- 0.3 per 2 h in the POA (P > 0.05). Melatonin (10(-8) M) did not alter the release of GnRH in the presence or absence of forskolin (10(-4) M). Melatonin, which was without effect on basal cAMP, inhibited forskolin-stimulated cAMP accumulation in the medium by 50% in the MBH and 40% in the POA. These results suggest that in our incubation system, melatonin does not modify GnRH release, but probably acts through the melatonin binding sites located in the hypothalamus to inhibit forskolin-stimulated cAMP.

Annual variations of in vitro GNRH release by hypothalamic explants in intact and castrated male mink: relations with LH, FSH and testosterone circulating serum levels.

In mink, a short-day breeder, testis growth begins in autumn (November), reaches a maximum in February, before matings occur, and decreases from March to very low volumes during spring and summer. To study the effects of season and testosterone feedback on gonadotrophin and GnRH secretion, the annual variations of LH, FSH, testosterone and GnRH were studied in intact and castrated mink. As portal blood sampling raised serious difficulties, an in vitro static incubation system was used for studying GnRH variations. In intact mink, serum LH concentrations did not vary significantly throughout the year, whereas FSH concentrations increased significantly between September and November then decreased to a minimum in January. Testosterone values rose significantly from November to a maximum from January to March, decreased very rapidly thereafter. Castration in November resulted in a significant increase in LH and FSH concentrations which remained higher than the values measured in intact males throughout the year. In long-term castrated mink, FSH concentrations did not fluctuate during the year, whereas LH concentrations showed an annual variation, with high values in April and August. For the study of in vitro GnRH liberation, medio-basal hypothalamic explants were incubated in Krebs-Ringer phosphate buffer for 3 periods of 15 min, and stimulated with copper complexed equimolarly with histidine (Cu/His, 200 microM) and prostaglandin E2 (PGE2, 10 microM). After Cu/His, the release of GnRH was 1 to 4 fold the basal release; after PGE2, the increase was 4-7 fold the basal release. The basal release of GnRH increased significantly between September and October to reach a maximum in November, decreased significantly in December to a minimum in February then increased progressively from May. The release of GnRH stimulated by Cu/His and PGE2 showed the same seasonal variation as the basal release. Castration 8 days before the sacrifice did not alter the release of GnRH, except in December: the release stimulated by PGE2 was then higher in intact than in castrated mink. Taken together, these results indicate that, with an in vitro static incubation system, it is possible to study the annual variations of GnRH release and to correlate these variations with those of serum gonadotrophin and testosterone concentrations. The synthesis and release of GnRH increased slightly from May, under long days, then more rapidly from September, resulting in an increased secretion of FSH in October, responsible for testis recrudescence. The annual pattern of basal and stimulated GnRH release was similar in intact and castrated mink, suggesting a direct effect of the season on the hypothalamus, rather than a negative feedback effect of the testis; however, testosterone seemed to feedback mainly at the pituitary level.

Influence of season of birth on onset of gonadotrophic and ovarian functions in young doe hares (Lepus europaeus).

The pituitary and ovarian responses to a monthly i.v. injection of 5 micrograms luteinizing-hormone-releasing hormone (LHRH) were studied in three groups of young doe hares, born in January-February (group I), in April (group II) or at the end of the breeding season (August-September, group III). The LHRH injection was always followed by a release of LH and progesterone, which did not differ among the three groups at 3 months of age. The pituitary and ovarian responses to LHRH increased gradually from the age of 3 months in groups I and III and from the age of 9 months in group II. One female of the ten born in January-February ovulated and reached puberty in June, at the age of 4 months, but with a weak pituitary response. The females born in April displayed a seasonally delayed puberty, at 9 months of age (two of five females ovulated in the next January). Four of the five females born at the end of the breeding season ovulated after LHRH when 5 months old (in February), with a full pituitary-ovarian response. The low pituitary response of group I in June-August, even if 10-20% of females ovulated after LHRH, suggests a need for a period of short days. Then, the most favourable conditions for the hare to reach puberty would be a period of short decreasing daylengths during the fall, followed by increasing daylengths after the winter solstice.

Gonadotrophins and sex steroids during pregnancy and natural superfoetation in captive brown hares (Lepus europaeus).

In brown hares, which are induced ovulators, sexual behaviour occurs episodically at the beginning of pregnancy. From Day 34 (length of pregnancy is 41 days), the frequency of sexual chases followed by mating, ovulation and fertilization increased and 59% of pregnant females presented a natural superfoetation. The pattern of circulating luteinizing hormone (LH), follicle-stimulating hormone (FSH), oestradiol and progesterone was studied in 13 pregnant females left permanently with a male, and in 10 females isolated from males around Day 20 of pregnancy. In the 2 groups FSH concentrations were high at the beginning and end of pregnancy. All females presented a peak value of FSH in the last 4 days of pregnancy, regardless of mating stimuli. This peak value was higher for females left permanently with a male than for isolated ones. Oestradiol concentrations fluctuated between 20 and 100 pg/ml, without any clear correlation with sexual behaviour, stage of pregnancy or profiles of other hormones. Prepartum matings occurred when progesterone values were still greater than 50 ng/ml; they were followed by a transient rise in LH and by a periovulatory progesterone secretion, with values above 100 ng/ml in the morning after mating. Such modifications of LH and progesterone were not detected before Day 34, suggesting that mating stimuli are not able to induce an LH surge at the beginning of pregnancy. After Day 34, mating can induce an LH surge, ovulation and superfoetation.

Pituitary and ovarian responses to luteinizing-hormone-releasing hormone during pregnancy and after parturition in brown hares (Lepus europaeus).

Female hares were given an i.v. injection of 5 micrograms luteinizing-hormone-releasing hormone (LHRH) between Days 7 and 19 (n = 21), 20 and 33 (n = 17) and 34 and 41 (n = 17) of pregnancy, and in the 3 days after parturition (n = 16). Whatever the stage of pregnancy, the LHRH injection induced a release of luteinizing hormone (LH) and follicle-stimulating hormone (FSH) and an acute secretion of progesterone; these hormonal responses increased significantly during pregnancy, to reach values similar to those observed in nonpregnant, nonpseudopregnant females during the breeding season in the 3 days after parturition. However, the release of LH remained monophasic in pregnant and post-partum females, in contrast to the unmated females during the reproductive season, in which there was a biphasic profile. The proportion of ovulating females after LHRH treatment was approximately 60% at the beginning and end of pregnancy; and, after parturition, fell to 23% between Days 20 and 33. After Day 33, the pituitary response to LHRH was significantly higher in ovulating than in nonovulating females. At the beginning of pregnancy, 67% of females aborted after LHRH injection; after Day 20, the incidence of abortion decreased significantly and was 0% from Day 34. The amplitude and duration of progesterone secretion by the new corpora lutea resulting from ovulation after LHRH injection were similar to those of corpora lutea induced in nonpregnant females during the breeding season.(ABSTRACT TRUNCATED AT 250 WORDS)

Effect of ovariectomy at two periods of the year on LH and FSH basal concentrations and pituitary response to LHRH in the brown hare (Lepus europaeus).

In the brown hare, fertile mating takes place from the beginning of December to September. Seasonal variations of basal concentrations of LH and FSH, and pituitary response to a monthly i.v. injection of LHRH were studied in intact control females and in females ovariectomized during the seasonal anoestrus (OVX1) or during the breeding season (OVX2). In intact females, both basal and LHRH-stimulated LH levels showed an annual variation, with minimal values during anoestrus. During the breeding season, the LH response to LHRH exhibited a biphasic pattern. In contrast, there was no clear seasonal variation in basal and LHRH-stimulated FSH concentrations. After ovariectomy during anoestrus, basal LH remained low for 2 months and began to increase in December. After ovariectomy during the breeding season, LH basal concentrations increased within a few days after the operation. Thereafter, LH values remained high in both groups of females until September, and decreased significantly as in intact females. The pattern of LH release after LHRH remained monophasic in the two groups of ovariectomized females. In OVX1 females, the LH response increased as early as October, was maximum from December to April and decreased progressively until October. IN OVX2 females, the LH response decreased regularly after ovariectomy to a minimum in October. In the 2 groups of ovariectomized females, basal FSH concentrations and pituitary response to LHRH rose rapidly after ovariectomy and did not vary significantly thereafter. These results showed a direct central effect of season on the regulation of basal concentrations of LH, modulated by a negative feed-back of ovarian secretions during the breeding season. In intact hares, the enhanced LH response after LHRH during the breeding season was related to an acute positive effect of ovarian secretions. The regulation of FSH was less dependent on season and remained under a negative control of the ovary throughout the year.

Pattern of serum concentrations of prolactin and progesterone during pregnancy and lactation in the brown hare (Lepus europaeus).

A heterologous radioimmunoassay system developed for the rabbit has been shown to measure prolactin in the hare. Concentrations of prolactin showed a significant (P less than 0.01) increase in the last 3 days of pregnancy (87.7 +/- 11.7 compared with 9.8 +/- 1.4 (S.E.M.) micrograms/l; n = 10 and 9 respectively) in pregnant females isolated from males, as well as in pregnant females kept with males and mating prepartum. This rise of prolactin at the end of pregnancy was not due to mating stimuli and occurred at a time when progesterone levels were still high (159 nmol/l). The injection of a slow-release preparation of bromocriptine (5 mg s.c.), which reduces prolactin secretion at the end of pregnancy, did not impair parturition. During lactation, prolactin levels increased significantly (61.2 +/- 19.8 compared with 5.3 +/- 0.1 micrograms/l; P less than 0.01) only after suckling stimuli.

Relative roles of oestradiol and of the uterus in the maintenance of the corpus luteum in the pseudopregnant brown hare (Lepus europaeus).

Brown hares were made pseudopregnant by sterile matings or PMSG-hCG treatment (day of mating or hCG injection = Day 0 of pseudopregnancy). Progesterone secretion by the CL began 3-4 days after the ovulatory stimuli, reached maximum on Days 8 to 11 and decreased thereafter to reach low levels from Day 9 to 18, depending on the female. Cauterization of all large ovarian follicles on Day 7 resulted in an immediate luteolysis in young females, but had no effect in older ones. Oestradiol capsules implanted from Day 7 to Day 46 were able to maintain progesterone secretion until at least Day 30, in intact females as well as in females with all large follicles cauterized. Hysterectomy on Day 7 or 8 was followed by an immediate drop in progesterone concentrations; oestradiol capsules implanted at the time of hysterectomy prevented the drop in progesterone values, which remained elevated until Day 38. The induction of ovulation in females hysterectomized 2 months before resulted in CL of slightly shortened life-span. The injection of PGF-2 alpha on Day 7 of pseudopregnancy was followed by an immediate luteolysis. These results suggest that oestradiol secreted by the large ovarian follicles is the main luteotrophic factor in the brown hare. In old hares, the large amount of interstitial tissue could secrete oestrogens, and thus maintain pseudopregnancy. On Day 7 of pseudopregnancy, the uterus secretes a luteotrophic substance acting either directly on the ovary, or via the pituitary, to maintain oestradiol secretion by the follicles. In long-term hysterectomized females, the CL would be able to develop independently of any trophic substance, but for a reduced duration.

Relation between circulating sex steroid concentrations and sexual behaviour during pregnancy and post partum in the domestic rabbit.

Sexual behaviour of 16 female and 12 male rabbits was studied during pregnancy and early post partum. The main behavioural events of the male (nuzzling and mounting) did not differ in the presence of receptive or non-receptive females. When introduced into the cage of the male, receptive and non-receptive females flattened to the floor or circled around. Sexual receptivity to males decreased in early pregnancy and increased to a maximum a few days around parturition; on Days 1 and 6 post partum, all experimental rabbits submitted to mating. Two groups of females were distinguished: one group submitting to mating whatever the stage of pregnancy, the second being receptive only during the few days before parturition or post partum. During pregnancy and early post partum there was no relation between the colour of the vulva and the female sexual behaviour. Does remained sexually receptive even when progesterone concentrations were high. Nevertheless, the number of receptive females was higher when progesterone concentrations decreased around parturition and the mean daily progesterone values were consistently higher in non-receptive than in receptive females. Oestrogen concentrations during pregnancy were very low and were not related to receptive behaviour.