Involvement of splenocytes in the control of corpus luteum function in the rat

We previously found that splenectomy caused a 1-day delay in ovulation in cycling rats. In the present study, the role of the spleen or splenocytes in corpus luteum function was investigated by comparing long-term splenectomized (SPX) rats with intact controls. When the rats were cervically stimulated on the day of proestrus, both SPX and control rats showed prolonged diestrus for 16 days associated with an increase in serum progesterone (P). However, in SPX rats, 20 alpha-dihydroprogesterone (20 alpha-OHP) concentrations were double those of controls throughout the period of pseudopregnancy. The concentration of total progestin (P plus 20 alpha-OHP) was also higher in SPX rats. All of these phenomena were normalized by injecting the animals with splenocytes. Taken together with our previous findings, these results indicate that splenocytes are involved in the control of ovarian function.     

Postcopulatory effects of two antifertility agents on ova transport and implantation

The daily doses which prevented implantation in 50 percent of treated animals (ED50) of 2,3 - bis (4-hydroxyphenol) valeronitrile (SC-3402) and 2,3 - bis (4-methoxyphenyl) pent-2-enenitrile (SC-3296) injected in rats s on Days 1 to 3, or Days 4 to 7, or Days 1 to 7 (Day 1 = pregnancy) were 100, 200, and 40 mcg and 50, 100, and 12 mcg respectively, ED50 doses of estrone were 4,8 and 3.5 mcg. Control animals showed ova in the oviduct only on Days 1, 2 and 3, also in the uterus on Day 4, and only in the uterus on Day 5. Very few ova were found in rats treated with 10 mcg estrone daily Day 1-2 and autopsied on Day 3. The same treatment period with 200 mcg SC-3402 caused similar results. 64 mcg SC-3402 resulted in a smaller reduction of ova. Acceleratory potency of 200 mcg SC-3402 is greater than can be due to its estrogenic activity equivalent, 0.5 mcg estrone; that of 64 mcg SC-3296 (4.8 equivalents estrone) can be so ascribed. Rats receiving daily 4-8 mg 17 alpha-acetoxy-6 alpha-methylprogesterone (MAP) from Day 1 to 9 to delay nidation, and 200 mcg SC-3402, autopsied on Day 10 showed no free blastocysts and a few implantation sites in the process of resorption (Free blastocysts were found in rats similarly treated but with ligation of the uterus at the cervix on Day 5 to prevent expulsion of blastocysts). Control rats on Day 10 showed a few implantation sites and free blastocysts. The normal number of implantations were present in SC-3296 treated rats. The average weight of cornu traumatized by threading one cornu in psuedopregnant rats with a silk thread on Day 5 (Day 1=cervical stimulus) in rats treated with 200 mcg SC-3402 on Days 5-8, 404 plus or minus 50 mg was significantly (P less than .05) lower than mean control weight, 794 plus or minus 48 mg. The difference between the mean weight of non-traumatized cornu of rats given 100 mcg, 284 plus or minus 36 and 232 plus or minus 12 mg respectively was significantly (P less than .05) greater than in controls, 159 plus or minus 5.8 mg. The deciduoma-inhibiting activity of SC-3402 is further evidence that it initiates nidation but impedes early implantation stages.     

Changes in ovarian blood flow and secretion of progesterone and 20 alpha-hydroxypregn-4-en-3-one on day 16 and the morning and afternoon of day 22 of pregnancy in the rat

During late pregnancy in rats, ovarian secretion of progesterone decreases and that of its reduced metabolite, 20 alpha-hydroxypregn-4-en-3-one (20 alpha-OHP), increases. The present study was undertaken to determine whether changes in ovarian blood flow are consistent with changes in progestin secretion. Rats (n = 5 per group) were examined on Day 16, the time of maximal progesterone secretion, and in the morning (AM) and afternoon (PM) of Day 22, the day prior to parturition. Ovarian blood flow was monitored continuously for 60 to 80 min, and serial samples of arterial and ovarian venous blood were obtained at 20-min intervals for determination of ovarian secretion rates of progesterone and 20 alpha-OHP. Ovarian blood flow increased from 0.38 +/- 0.04 ml/min (mean +/- SEM) on Day 16, to 0.77 +/- 0.05 and 0.78 +/- 0.04 ml/min on Day 22 AM and PM, respectively, whereas the secretion of progesterone decreased from 26.9 +/- 4.0 to 4.5 +/- 1.0 and 3.2 +/- 0.3 micrograms/h per ovary. The secretion of 20 alpha-OHP was similar on Day 16 and Day 22 AM (5.6 +/- 1.7 and 5.4 +/- 1.3 micrograms/h per ovary) but then increased to 18.9 +/- 1.2 micrograms/h per ovary by Day 22 PM. Thus the amount of total progestins secreted per unit rate of blood flow relative to that on Day 16 (100%) fell to 15% and 34% on the morning and afternoon of Day 22, respectively. Clearly, the relative changes in ovarian progestin secretion and blood flow in the rat near term to not conform to patterns observed at luteal regression in some other species.(ABSTRACT TRUNCATED AT 250 WORDS)     

Splenic macrophages enhance prolactin and luteinizing hormone action in rat luteal cell cultures.

We have reported that splenic macrophages play a role in the regulation of progestin secretion in rats. In this study, splenic macrophages were obtained from cycling rats at different estrous cycle stages and co-cultured with luteal cells from mid-pseudopregnant rats in the absence/presence of prolactin (PRL) or luteinizing hormone (LH). The effect of macrophages on the luteotropic action of PRL and LH was evaluated with 2 parameters, i.e. an increase in total progestin output (progesterone plus 20 alpha-hydroxyprgn-4-en-one [20 alpha-OHP]), and an increase in the progesterone to 20 alpha-OHP (P/20 alpha-OHP) secretion ratio. Splenic macrophages obtained from proestrous or metestrous rats enhanced the PRL action to increase the P/20 alpha-OHP secretion ratio, but those from estrous or diestrous donors did not. Only macrophages from proestrous donors enhanced the PRL action to increase the total progestin output. In contrast, the LH action increasing the P/20 alpha-OHP secretion ratio was enhanced by splenic macrophages regardless of the donors' estrous cycle stages. The LH action increasing the total progestin output was enhanced only by proestrous or metestrous macrophages. Therefore, if luteal cells are co-cultured with proestrous macrophages, the luteotropic actions of PRL and LH can be fully expressed. These results indicate that splenic macrophages directly act on luteal cells and enhance the luteotropic action of PRL and LH, and that this function of splenic macrophages is modified somehow according to the donors' estrous cycle stages.     

Quantitative changes in steroidogenic organelles in the corpus luteum of the pregnant rat in relation to progestin secretion on day 16 and in the morning and afternoon of day 22

The present study was carried out to determine whether the major steroidogenic organelles of luteal cells quantitatively reflect variations in ovarian steroid secretion rates during pregnancy in the rat. Assessments were made on day 16 and in the morning (AM) and afternoon (PM) of day 22 (term is day 23). Ovarian steroidogenesis differs both quantitatively and qualitatively between these stages of pregnancy, so together they provide an ideal physiological model to study structural-functional relationships in the ovary. Corpora lutea of five rats were examined at each stage after progesterone and 20 alpha-hydroxypregn-4-en-3-one (20 alpha-OHP) secretion rates had been determined by a venous outflow technique. Total progestin secretion (progesterone plus 20 alpha-OHP) fell from 32.5 +/- 5.2 to 9.8 +/- 1.2 micrograms/hr per ovary (mean +/- SEM) between day 16 and day 22 AM but then increased to 22.6 +/- 1.4 micrograms/hr per ovary by day 22 PM. The total volume of luteal cell cytoplasm was slightly greater at day 22 AM and PM than at day 16. Similarly, the volumes of smooth endoplasmic reticulum (SER), lipid droplets, and membrane-bound granules all increased, but the volume of mitochondria decreased slightly. In contrast, the surface areas of SER and the inner and outer mitochondrial membranes did not change between day 16 and day 22 AM but then increased substantially by day 22 PM. Therefore, steroid secretion rates per unit area of steroidogenic membrane showed no consistent pattern over the stages examined.(ABSTRACT TRUNCATED AT 250 WORDS)     

Levels of prostaglandin F (PGF) in bovine endometrium uterine venous, ovarian arterial and jugular plasma during the estrous cycle (38789).

Prostaglandins F (PGF) were measured in uterine vein, ovarian artery, and jugular vein plasma and in the endometrial tissues at various times during the bovine estrous cycle, and were compared to peripheral plasma progesterone levels. Four groups of heifers at days 1-5, 10-14, 15-17 and 20-0 of the estrous cycle were studied. Low levels of PGF (48 plus or minus 12 ng/g dry tissue were measured in the endometrium on days 1-14 of the cycle. Higher values (131 plus or minus 9.0) were found at days 15 until the day of estrus (p less than 0.001). Similarly, very low levels of PGF were observed in the uterine vein plasma at days 1-14 (0.162) plus or minus 0.044) ng/mlM plus or minus S.E.), whereas on days 15 until the day of estrus the levels ranged from 1.5 to 3.0 ng/ml. The increases in uterine vein PGF on day 15 occurred even while peripheral plasma progesterone levels were still high. However, PGF was not elevated in either the ovarian artery or the jugular vein at any time during the cycle, even when uterine vein levels were greatly elevated. No differences in PGF content were detected in endometrial tissue from uterine horns adjacent or opposite to the functional corpus luteum.     

Midtrimester pregnancy interruption and the placental progesterone levels.

Placental progesterone contents were studied in 10 patients therapeutically aborted at midtrimester by intraamniotic infusions of hypertonic saline, and from 14 patients (12-20 weeks) aborted by intraamniotic instillation of prostaglandin F-2alpha. The mean S.E. of the progesterone was 1.99 plus or minus 0.07 ug/g. placental tissue in the first group, while with prostaglandin abortion the placental progesterone was 1.45 plus or minus 0.08 ug/g. tissue, which is significantly lower than the results in the first group (P equals 0.001). The possible mechanism of action of prostaglandin as an effective abortifacient is discussed.     

Ovarian responses to perphenazine-induced prolactin secretion in the rats: Effect of ovulation, stress, and steroids.

A single injection of 2.5 mg perphenazine (PH)/kg body wt to rats on the day of estrus (day 0) did not result in increased serum progesterone 24 hr later. Continued daily injections, however, resulted in a 2.5-fold increase in serum progesterone between days 1 and 3 and a 1.6-fold increase between days 3 and 5 to a final concentration of 58 plus or minus 4 ng/ml on day 5 in serially anesthetized and bled rats. Neither daily administration of 5.0 nor 10.0 mg PH/kg body wt to rats subjected to the stressful conditions of this regimen resulted in further increases in serum progesterone, but the 5.0 mg dose of PH in unstressed rats bled only on day 5 resulted in a highly significant increase in serum progesterone to 110 plus or minus 7 ng/ml. In unstressed rats the increase in serum progesterone over control values after five daily injections of 2.5 mg PH/kg body wt could be attributed to decreased 20alpha-reduction of progesterone, but when the dose of PH was increased to 5.0 mg/kg, a highly significant increase in both progesterone and total progestins occurred indicating that prolactin can increase steroidogenesis as well as reduce 20alpha-hydroxysteroid dehydrogenase activity. After inhibition of ovulation, the 5.0 mg daily dose of PH resulted in serum progesterone of only 25 plus or minus 8 ng/ml on day 5 in unstressed rats. Thus, serum progesterone in ovulating rats treated with PH originated primarily in the corpora lutea. Perphenazine, 5.0 mg/kg, administered only on estrus and the first day of diestrus was sufficient to induce pseudopregnancy of 14.5 plus or minus 1.6 days. No evidence for gonadotropin stimulation of the ovaries of any rats was observed. The effect of stress on the progesterone response was not mimicked by administration of cortisol acetate and is assumed to be medicated by suppression of prolactin secretion.     

Endocrine control of clutch size in reptiles. VIII. antiestrogenic effects of clomiphene citrate in the lizard Anolis carolinensis

The influence of clomiphene citrate on follicle-stimulating-hormone (FSH) and estradiol-induced growth of ovarian follicles and oviducts in the lizard A. carolinensis was studies. In Experiment 1 lizards received 14 daily injections of either saline, clomiphene (1, 10 or 20 mcg), or FSH (1 or 10 mcg) or combined clomiphene-FSH treatment. In Experiment 2, adult lizards with hypertrophied, vitellogenic ovaries, and enlarged oviducts, weres adenohypophysectomized and treated with a daily dose of .05 ml of either saline, saline plus 5 mcg clomiphene, saline plus 10 mcg FSH, saline plus 10 mcg estradiol-17beta, or FSH plus clomiphene or estradiol plus clomiphene. FSH increased follicle size in previtellogenic ovaries. Injection of 1 mcg clomiphene reduces the effects of FSH. 20 mcg clomiphene given alone stimulated the growth of larger follicles. Clomiphene blocked FSH-induced appearance or maintenance of large (less than 2.0 mm) vitellogenic follicles. It blocked FSH gains in oviductal weight and well as stimulated growth of small previtellic follicles. Estradiol-induced follicular and oviductal growth was uneffected by clomiphene. While low doses of clomiphene are antiestrogenic they are unable to combat the effects of high dose estradiol.     

The effects of cervical dilation on plasma PGFM, progesterone and the duration of luteal function in diestrous mares

Transcervical diagnostic techniques may alter the length of the equine estrous cycle and affect subsequent luteal function. Therefore, nine mares were used to determine the effect of cervical dilation on plasma 13, 14-dihydro, 15-keto-prostaglandin F(2) (PGFM), progesterone (P(4)) and posttreatment duration of luteal function. Mares were given a daily score of 0 to 4 based on sexual receptivity. Five days following the end of receptivity, mares were randomly assigned to one of three, 3 x 3 latin squares. Control mares received no cervical dilation. Cervically stimulated mares recieved cervical dilation for 60 sec. Cervically stimulated plus inhibitor mares were dilated similarly to cervically stimulated mares, but received a prostaglandin synthetase inhibitor 30 min prior to treatment. Each mare completed all three treatments in three consecutive estrous cycles. Plasma PGFM and P(4) were determined by RIA. Plasma PGFM was lower (P<0.05) in cervically stimulated plus inhibitor than control and cervically stimulated mares. In addition, plasma P(4) was lower (P<0.10) in cervically stimulated plus inhibitor than in control and cervically stimulated mares. Luteal function following treatments did not differ. These data indicate that neither plasma PGFM and P(4) nor the duration of luteal function were affected by cervical dilation. However, administration of a prostaglandin synthetase inhibitor prior to cervical dilation decreased plasma PGFM and P(4) concentrations.     

Luteolysis in the hamster: abrogation by gonadotropin and prolactin pretreatment

The luteolysis which terminated pseudopregnancy (PSP) in superovulated hamsters was studied. Spontaneous luteolysis occurred before 1100 on Day 7 of PSP and was characterized by a rapid decline in circulating progesterone levels. Luteolysis induced by prostaglandin F2 alpha (PGF2 alpha) on Day 5 of PSP displayed a similar rapid reduction in progesterone over 24 hours. In both cases levels of the progesterone metabolite 20 alpha hydroxypregn-4-ene-3-one (20 alpha-OHP) were less than 2 percent of progesterone levels and declined in a manner similar to progesterone. This suggests that conversion of progesterone or its precursors to 20 alpha-OHP was not a functional aspect of luteolysis in the hamster. Pretreatment with either prolactin (PRL), luteinizing hormone (LH) or follicle stimulating hormone (FSH) failed to prevent PGF2 alpha-induced luteolysis on Day 5 in the superovulated PSP hamster. Combinations of PRL and LH, LH and FSH or PRL and FSH were also unsuccessful in abrogating luteolysis. However, pretreatment with a combination of PRL, FSH and LH prevented luteolysis in 11/14 animals. These results suggest that luteotropic agents can reverse the luteolytic effects of PGF2 alpha in the hamster.     

Adrenergic activity in hypothalamus and ovulation

The turnover of norepinephrine (NE) and its synthesis in the hypothalamus in rats just prior to and subsequent to ovulation was studied. Hypothalamic NE concentration in proestrus (preovulatory) was higher than in estrus rats (3.48 plus or minus) .09 mcg/gm vs. 2.13 plus or minus .04 mcg/gm). When methylester hydrochloride (MPT), a blocker of catecholamine biosynthesis, was injected, proestrus rats NE dropped 62% vs. estrus rats' drop of 28.6%. Tritiated NE injected to show synthesis rates showed a higher rate of NE synthesis produced in the hypothalamus during proestrus vs. estrus. In addition there was an increase in NE levels between diestrus Day 2 and proestrus localized to the anterior and middle hypothalamus.     

Qualitative and quantitative data on experimental luteinization in the female rat

The dose-effect of 1.5-16 mcg luteinizing hormone (LH) per 100 gm body weight injected in rats at 1100-1200 on proestrus was compared with 30 mg meprobamate given to controls at the same time, on luteinization and ovulation seen in serial ovarian sections. The WII Wistar rats were killed. Luteinization with or without ovulation increased with dose (1.5, 3, 5, 8, and 16 mcg) of LH to a plateau (90%) above the 5 mcg dose, compared with 18% in controls. 2-5 animals in each dose group had preluteinized follicles, characterized by a dissociation of the cumulus oophorus from the granulosa. The absolute frequency of ovulation increased linearly with LH dose, but the frequency of ovulation among rats that luteinized was invariant. The coefficient of ovulation, calculated as the mean incidence of ovulation in relation to the total number of luteinized or preluteinized follicles in each rat, decreased from .769 in controls to .580 in the 3 mcg group, then rose to .916 in the 16 mcg group. Thus, in proestrous rats, low doses of LH induce corpora lutea with retained ova. The threshold dose of LH for luteinization and for ovulation is lower in proestrus than in diestrus II, but varies slightly in different strains of rats.     

Physiologic role of relaxin on mammary gland growth in rats

The role of relaxin in stimulating growth of the mammary gland was assessed in ovariectomized and intact male rats for a period of 20 days. In addition to relaxin alone, the ovarian mammogenic hormones estradiol and progesterone were used in combination with relaxin and with each other to evaluate responses of mammae. Indices for mammary growth included wet weight, dry fat-free tissue, DNA, RNA, total protein, and collagen. Quantitative estimates of DNA and collagen represented the best indicators of parenchymal and stromal growth, respectively. Because changes in body weights were significantly different among hormonally administered groups, these were included as well. In Ovariectomized young rats, relaxin alone and in combination with estradiol and progesterone increased all indices significantly (P less than 0.01). The collagenous portion of total protein was high for the group receiving relaxin alone (62%) compared with the control group (46%). Relaxin administered along with estradiol and progesterone increased collagen accumulation to 73%, compared with 54% in the estradiol + progesterone group. Relaxin did not significantly increase growth indices when administered to male rats at 10 and 20 micrograms/day, while 30 micrograms stimulated a significant increase in total protein (P less than 0.05), suggesting that 30 micrograms of relaxin/day may be considered the basal concentration needed to induce a physiologic response in males. Relaxin induced a growth effect on mammae by synergizing with progesterone and estradiol in order to stimulate parenchymal proliferation, as noted by a DNA increase, and to increase stromal distensibility of the mammary pad by invoking accumulation of collagen and total protein in substituting for mammary adipose tissue.     

The non-genomic effects on Na+/H+-exchange 1 by progesterone and 20alpha-hydroxyprogesterone in human T cells

Progesterone is an endogenous immunomodulator and can suppress T-cell activation during pregnancy. We have previously shown that the non-genomic effects of progesterone, especially acidification, are exerted via plasma membrane sites and suppress cellular genomic responses to mitogens. This study aimed to show that acidification is due to a non-genomic inhibition of Na(+)/H(+)-exchange 1 (NHE1) by progesterone and correlate this with immunosuppressive phytohemagglutinin (PHA)-induced T-cell proliferation. The presence of amiloride-sensitive NHE 1 was identified in T cells. The activity of NHE1 was inhibited by progesterone but not by 20alpha-hydroxyprogesterone (20alpha-OHP). Furthermore, 20alpha-OHP was able to compete with progesterone and release the inhibitory effect on the NHE1. The inhibition of NHE1 activity by progesterone-BSA demonstrated non-genomic action via plasma membrane sites. Finally, co-stimulation with PHA and progesterone or amiloride, (5-(N, N-dimethyl)-amiloride, DMA), inhibited PHA-induced T-cell proliferation, but this inhibition did not occur with 20alpha-OHP and PHA co-stimulation. However, when DMA was applied 72 h after PHA stimulation, it was able to suppress PHA-induced T-cell proliferation. This is the first study to show that progesterone causes a rapid non-genomic inhibition of plasma membrane NHE1 activity in T cells within minutes which is released by 20alpha-OHP. The inhibition of NHE1 leads to immunosuppressive T-cell proliferation and suggests that progesterone might exert a major rapid non-genomic suppressive effect on NHE1 activity at the maternal-fetal interface in vivo and that 20alpha-OHP may possibly be able to quickly release the suppression when T cells circulated away from the interface.     

Multiple inhibitory effects of a luteinizing hormone-releasing hormone agonist on hCG-dependent steroidogenesis and on FSH-dependent responses in ovarian cells in vivo and in vitro

[125I] labelled [D-Leu6, des-Gly-NH10(2)] LH-RH ethylamide (LH-RHa), when injected into immature female rats, bound specifically not only to the pituitary but also to the ovaries. LH-RHa inhibited hCG-stimulated progesterone production and ovarian weight augmentation in hypophysectomized immature female rats in vivo. FSH-induced ovarian hCG receptors and ovarian weight gain in diethylstilbestrol (DES)-treated hypophysectomized immature female rats were also suppressed by LH-RHa. Progesterone production by rat luteal cells in vitro was inhibited by LH-RHa. LH-RHa did not change the affinity or population of LH/hCG receptor in porcine granulosa cells in short term incubation. However, LH-RHa inhibited induction of LH/hCG receptor stimulated by FSH and insulin in long term culture of porcine granulosa cells. LH-RHa delayed hCG-stimulated cyclic AMP accumulation in porcine granulosa cells. These findings suggest that LH-RHa inhibits hCG-stimulated cyclic AMP accumulation and subsequent progesterone production as well as FSH-stimulated LH/hCG receptor induction by acting directly on ovarian cells.     

Maintenance of unimplanted fertilized ova in sprayed rats. IV. Effects of duration of ovarian hormone deprivation and progesterone therapy prior to the deprivation

Studies were carried out in the spayed-rat delayed-implantation model to determine whether progesterone treatment prior to an ovarian hormone deprivation during the pre-implantation period would influence the incidence of subsequent delayed ovo-implantation induced with progesterone plus estrone. Implantation was rarely induced with 4 mg progesterone plus 1 microgram estrone/day after 5 to 11 days of ovarian hormone deprivation in rats that were spayed on Day 3, if progesterone treatment were not given before ovarian hormone deprivation. In contrast to this, implantation was fairly consistently induced with 4 mg progesterone plus 1 microgram estrone/day after 3, 5, 7, or 11 days of ovarian hormone deprivation in rats that were spayed on Day 3 and received 4 mg progesterone/day before the deprivation period (i.e., on Days 2 through 3, 2 through 6, or 2 through 8). The post-implantation viability of the embryos of the dams undergoing the longer periods of ovarian hormone deprivation, however, was reduced.     

Gonadotropin-releasing hormone (GnRH) inhibits ovulation induced with luteinizing hormone (LH) in proestrous hypophysectomized rats

In this paper we present evidence that a single low dose of the natural synthetic gonadotropin-releasing hormone (GnRH), inhibits ovulation induced by LH in proestrous-hypophysectomized rats. Rats hypophysectomized by the parapharyngeal route in the morning of proestrus received an intravenous injection of 100 or 300 ng GnRH at 1400 h immediately followed by 1.0 microgram LH per 100 g bw. In control groups, either one or both hormones were replaced with 0.9% NaCl. Ovulation was assessed the following morning by counting the ova present in oviductal flushings. All the rats treated with LH alone ovulated, and the addition of GnRH reduced significantly the number of ovulating rats and the number of ova per ovulating rat. In other groups of rats hypophysectomized in the morning of proestrus and treated in the same way, ovarian or adrenal secretory rates of estradiol and/or progesterone were measured after cannulation of the corresponding vein, in the afternoon of proestrus. In these animals, GnRH failed to inhibit either the ovarian progesterone surge observed 2 h after LH administration, or the adrenal progesterone secretion. All hypophysectomized rats showed lower ovarian secretory rate of estradiol than intact rats; this rate was not affected by treatment with LH or LH plus GnRH. The systemic estradiol levels in plasma of hypophysectomized rats were distributed within a range of 20 pg/ml to 50 pg/ml. The number of rats whose levels were above 21 pg/ml on estrus day was significantly higher in rats receiving 300 ng GnRH as compared to those receiving 100 ng GnRH, reaching values that surpassed the concentration found in intact, untreated animals at the same time of estrus. This effect did not depend on LH administration.