Plasma and pituitary concentrations of LH, FSH and prolactin after injection of GnRH in aged female C57BL/6 mice.

Plasma and pituitary concentrations of LH, FSH and prolactin (Prl) were compared by radioimmunoassay (RIA) in young mature (2-4 month-old) and aged (16-22 month-old) female C57BL/6 mice, following various experiments with IP injections of GnRH. Unanesthetized mice were bled by cardiac puncture either (1) at various time intervals following a single injection of 25 ng GnRH, (2) after several injections of 25 ng GnRH, or (3) after receiving excessively large doses of GnRH. From the RIA of plasma and pituitary concentrations of LH, it appeared that the pituitaries of aged mice were, in general, capable of responding to GnRH treatment, only to a lesser degree than that of the younger females. The exception was that the pituitaries of aged mice that were subjected to unusually high doses of GnRH continued to secrete higher levels of LH, whereas the response of the pituitaries in younger mice plateaued after receiving 25 ng GnRH. Plasma and pituitary concentrations of FSH and Prl remained fairly constant when aged GnRH-treated females were compared with aged controls. This was also true when FSH and Prl concentrations in the younger mice were compared with their controls, except for significantly higher plasma FSH and significantly lower pituitary FSH following several injections of GnRH. When the hormonal concentrations of the young and aged groups were compared in the various experiments, the older mice consistently exhibited higher levels of gonadotropin and lower levels of prolactin than those of the younger mice. In a separate experiment, in which a radioreceptor assay (RRA) was used to quantitate plasma LH in GnRH-treated mice, both age groups were found to have lower concentrations of LH (p less than 0.05) than those obtained by RIA from the same animals. The RRA to RIA ratio, however, was 0.84 for 2-4 month-old and 0.57 for 16-22 month-old mice. These studies suggest that the anterior pituitary of the aged mouse is less sensitive to GnRH stimulation.

Plasma and pituitary concentrations of LH, FSH, and prolactin in aging C57BL/6 mice at various times of the estrous cycle.

Plasma and pituitary concentrations of LH, FSH, and prolactin (Prl) were measured by RIA in 2-4, 7-8, 12-13 and 16-20 month-old female C57BL/6 mice during various stages of the estrous cycle. In general, gonadotropin concentrations tended to rise with increasing age and Prl concentrations tended to decline. Pronounced differences existed, however, between the four age groups around the time of the LH surge. LH secretion declined progressively with increasing age at 21.00 hr of proestrus. Aged mice, 16-20 months old, had significantly lower plasma concentrations of LH than did other age groups. It is not known whether age-related changes in the ovary, pituitary, or hypothalamus are largely responsible for differences in the secretion of LH, FSH and Prl in aging C57BL/6 mice.

Exposure to estradiol impairs luteinizing hormone function during aging.

The loss of ovulatory cyclicity in many mammals is caused by changes in the hypothalamic-pituitary (H-P) control of the preovulatory luteinizing hormone (LH) surge. This work evaluated the anterior pituitary (AP) component of the H-P axis by determining the ability of perifused AP to release LH following sustained but pulsatile LHRH stimulation. The normal dual discharge profile of LH was affected by age. The first hour of the response, unaffected by cycloheximide, was similar in 5-6-month-old (mature), 12-13-month-old (declining litters) and 16-18-month-old (irregularly cycling) mice. The remaining protein synthesis-dependent part of the response was reduced in the 16-18 and 22-24-month-old (anestrus) mice. The role of estradiol (E2) in AP aging was further tested as AP from ovariectomized (OVXed) mice, deprived of E2 since puberty, responded as well as the mature proestrous group. In contrast, aged mice subjected to long-term E2 exposure (cycling or OVXed plus E2 replacement) failed to produce the dual response pattern. Since alterations in LH response occurred during the protein-dependent phase, synthetic processes that involve packaging and transport of stored LH, or the production of new LH, may be affected by age. Furthermore, E2 is a major factor in altering LH function and appears to act before middle age.

Luteinizing hormone levels in aged female laboratory rodents as measured by a radioimmunoassay and a radioreceptor assay.

Plasma levels of LH were measured in young sexually mature (2- to 5-month-old) and aged (13- to 20-month-old) female C57BL/6 mice, Syrian hamsters and Wistar rats using a radioimmunoassay (RIA) and a radioreceptor assay (RRA). There were no statistically significant differences when comparing data from the two assays when examining young oestrous and dioestrous mice. Aged oestrous and dioestrous mice exhibited significantly higher levels of LH as measured by RIA than by RRA. Levels of LH analysed by RIA were also higher in aged mice compared with those in younger mice. Comparing LH hamsters produced similar results. In contrasts, aged pseudopregnant rats exhibited significantly lower levels of plasma LH than younger dioestrous females and there were no differences in RIA and RRA values. There were also no differences when comparing data from the two assays in younger rats. The mean (+/- S.E.M.) RRA: RIA ratios of 0.96 +/- 0.05 (oestrous) and 0.96 +/- 0.04 (dioestrous) for younger mice and 0.92 +/- 0.08 for younger hamsters were significantly higher than the ratios of 0.66 +/- 0.05, 0.72 +/-0.05 and 0.71 +/- 0.05 respectively of their aged counterparts. The mean RRA: RIA ratios in the two age groups of rats were almost identical (0.79 +/- 0.05 and 0.80 +/- 0.10). These studies suggested that a portion of the higher LH levels detectable by RIA in some aged female rodents results from qualitative changes in the LH molecule.

Neuroendocrine studies in hyperprolactinaemic male mice.

In several species, including man and the rat, hyperprolactinaemia is associated with suppression of gonadotrophin release and male sexual behaviour. However, in the hyperprolactinaemic male mouse, plasma LH and FSH levels and copulatory behaviour are increased rather than suppressed. In an attempt to identify mechanism(s) which may be responsible for these effects of hyperprolactinaemia in the mouse, we have examined the effects of two ectopic pituitary isografts on several indices of hypothalamic and pituitary function in adult DBA/2J males. Animals with pituitary grafts had markedly increased plasma concentrations of prolactin, LH and FSH and enlarged seminal vesicles, whereas testicular and pituitary weights were not affected. Content of LHRH receptors and activity of aromatase in the pituitary, as well as dopamine-beta-hydroxylase activity in the hypothalamus were nearly identical in pituitary-grafted and sham-operated males. Biosynthesis of dopamine and turnover of noradrenaline in the median eminence were significantly increased in grafted males. We suggest that the increase in the activity of hypothalamic noradrenergic neurones may mediate stimulatory action of hyperprolactinaemia on LH and FSH release in the mouse. Comparison of these results with those obtained previously in the rat suggests that species differences in the effects of prolactin on gonadotrophin release may be related to its divergent effects on noradrenaline turnover.

Plasma and pituitary concentrations of LH, FSH and prolactin in aging Mongolian gerbils.

Samples of plasma and pituitary homogenates collected from female Mongolian gerbils (3-4, 11-13 and 20-25 month-old) at various stages of the estrous cycle were analyzed by radioimmunoassay for luteinizing hormone (LH), follicle-stimulating hormone (FSH) and prolactin (Prl). Plasma and pituitary LH concentrations were similar in all three age groups. Plasma FSH concentrations tended to increase with age while plasma Prl concentrations remained unchanged. Pituitary concentrations of FSH and Prl were variable between the age groups depending upon the stage of the estrous cycle. The inability of older gerbils to produce young appears to result primarily from age-related changes occurring in the uterus rather than from alterations in the hypothalamic-hypophyseal complex.

Concentrations of prostaglandins in plasma, seminal vesicles, and ovaries of aging C57BL/6NNia mice.

Concentrations of prostaglandin E1 (PGE1), and prostaglandin E2 (PGE2) (combined in the same radioimmunoassay) and prostaglandin F2 alpha (PGF2 alpha) were analyzed in circulating plasma and seminal vesicles of 3- and 26 to 27-month-old males and in circulating plasma and ovaries of 3-, 6-, 14 to 18- and 26 to 30-month-old female C57BL/6NNia mice. The amount of PGE declined in the plasma (P less than 0.05) and seminal vesicles (P less than 0.02) of aged male mice, whereas PGF2 alpha concentrations remained unchanged. There were no statistical differences in plasma or ovarian concentrations of PGE or PGF2 alpha when comparing the various age groups of female mice. It does not appear as if age-related changes in prostaglandins play a significant role in reproductive senescence.

Plasma and pituitary concentrations of luteinizing hormone, follicle-stimulating hormone and prolactin in aged, ovariectomized CD-1 and C57BL/6 mice.

Plasma and pituitary concentrations of luteinizing hormone (LH), follicle-stimulating hormone (FSH), and prolactin (Prl) were determined by radioimmunoassay in young (2-4 months-old) and aged CD-1 (14-18 months-old) and C57BL/6 (16-22 months-old) mice one month after ovariectomy. In young, ovariectomized mice, plasma and pituitary concentrations of LH and FSH were significantly higher, whereas concentrations of Prl were significantly lower than those in control mice (sham-operated). In contrast, plasma concentrations of LH, FSH and Prl were not statistically different in aged, ovariectomized mice and aged, control mice. There were also no differences in pituitary concentrations of the three hormones when comparing the same aged C57BL/6 mice, although the aged, ovariectomized CD-1 mice exhibited higher pituitary levels of each hormone than those of their controls. The pituitary of the aged mouse responds differently to ovariectomy than that of the young mouse because of age-related changes in the ovary and/or hypothalamic-hypophyseal complex.

Orthotopic ovarian transplantation in young and aged CBA mice.

Reproductive and hormonal changes associated with CBA mice which were orthotopically transplanted with ovaries from young or aged CBA mice were studied. Reproductive decline was defined by the number of mice mating and the number of implantation sites and resorptions in pregnant mice. A high percentage of resorptions was observed in aged mice receiving ovaries from young mice. An increase in the number of resorptions and a decrease in the number of implantation sites was observed in young mice transplanted with aged ovaries. Concentrations of LH, FSH and Prl were analyzed by radioimmunoassay from plasma of pregnant and nonpregnant transplanted mice, ovariectomized and sham-operated controls. Young ovariectomized mice had elevated concentrations of LH and FSH when compared to sham-operated controls. No differences were noted between the aged ovariectomized mice although the gonadotrophin levels in these animals were higher than those in comparable young groups. The highest LH and lowest FSH concentrations were found in young to young transplanted mice. The Prl concentrations in transplanted mice did not vary between groups. These data suggest that the hypothalamic-hypophyseal complex is functioning adequately in all transplanted groups and age-related changes in the ovary and uterus are responsible for embryonic mortality.

GnRH receptor site increase on the surface of cultured gonadotropes of senescent C57BL/6NNia mice.

Anterior pituitary cells were cultured for 2 days from 6-, 14-, and 23-25-month-old C57BL/6NNia mice. The cells were then stimulated with one of three biotinylated GnRHs [biotinyl-Lys6]-[D-Lys6]GnRH, [biotinyl-Ser4]-[D-Lys6]GnRH, [biotinyl-Ser4]-[D-Trp6, des-Gly10]GnRH) at 4 degrees C for 1 h. Some of these cells were processed unfixed, attaching avidin-fluorescein and examined with fluorescent microscopy. Other cells were fixed and the biotinylated GnRH coupled to avidin-gold (20 nm particles), which was subsequently silver-enhanced (70 nm particles) and examined with light and scanning electron microscopy (SEM). To enhance the location of receptor sites for quantitation, a back-scattering electron detector was employed with the SEM. Gonadotropes from all three age groups ranged in size from 3 to 13 microns in diameter. The largest gonadotropes in each age group displayed the highest number of GnRH receptors, but the concentration/surface area (microns2) was less than for smaller cells. Smaller gonadotropes (3-6 microns diameter) from 14- and 23-25-month-old mice contained more GnRH receptor sites/microns2 (p < .001) than those from 6-month-old mice; the percentage of these gonadotropes was markedly higher for 14-(65%) and 25-(52%) versus 6-month-old (5%) mice. The number of receptor sites in larger gonadotropes (6.1-13 microns diameter) did not decline with the increasing age of the mouse. Decreased secretory activity in gonadotropes of 25-month-old mice may result from age-related changes in pituitary peptide processing or release, but it is not due to a lack of GnRH receptors or the inability of GnRH to bind to its receptor site.

Effect of ovariectomy and estrogen replacement on hypothalamic, pituitary and peripheral blood beta-endorphin levels in the rat.

This study was initiated to detect possible changes in beta-endorphin (beta-EP) levels of the hypothalamus, anterior pituitary gland, and peripheral blood of rats after ovariectomy and estrogen administration. Attempts were also made to determine the correlation between peripheral and central levels of beta-EP. Twenty-six Sprague-Dawley rats were decapitated. Nine had intact ovaries (Gr. INT), and 17 were ovariectomized 3 weeks before they were killed. Nine of the ovariectomized rats received estradiol benzoate (EB) (Gr. EB) and the other 8 received peanut oil (Gr. OVX) prior to the decapitation. A beta-EP radioimmunoassay was used to analyze homogenates of the hypothalamus and anterior pituitary, and peripheral blood. In the hypothalamus, beta-EP levels were significantly lower in Gr. INT and Gr. EB than in Gr. OVX. In the pituitary gland and peripheral blood, beta-EP levels were significantly higher in Gr. INT than in Gr. OVX. Pituitary beta-EP levels did not vary between Gr. OVX and Gr. EB, although beta-EP levels in peripheral blood were significantly higher in Gr. EB than in Gr. OVX. No significant correlations were noted in beta-EP levels between the hypothalamus, pituitary gland, and peripheral blood in either Gr. INT, Gr. OVX, or Gr. EB. It appears that EB exerts different effects on beta-EP levels in the hypothalamus, anterior pituitary gland, and peripheral blood, and that beta-EP levels in these regions may be independent of one another.

Acute lithium treatment suppresses the proestrous LH surge in mice: chronic lithium leads to constant diestrus.

Although the therapeutic usefulness of lithium in manic-depressive psychosis is now well-established, a number of basic and clinical studies in recent years have shown that the administration of this anti-manic drug produces a wide range of adverse endocrine and metabolic effects. The present study was undertaken in order to examine (a) what effects acute lithium administration might have on the preovulatory surge of luteinizing hormone (LH) during proestrus, and (b) whether chronic lithium administration has any adverse effect on the estrous cycle in C57BL/6 mice. Acute injections of lithium on the day of proestrus (at 10.00, 16.00 and 18.00 h; LD 14:10; lights on at 05.00 h CST) at a dosage of 5 mEq/kg b. wt. led to a significant (P less than 0.01) suppression of the LH surge that normally occurs in the evening of proestrus at 21.00 h. Chronic administration of lithium, on the other hand, resulted in a complete disruption in the regularity of the estrous cycle. This was characterized by an increasing number of mice showing a continuous diestrous vaginal smear during the first week of exposure to lithium, after which all of the lithium-treated mice completely stopped cycling and entered into constant diestrus. These results represent for the first time that lithium has significant adverse effects on the reproductive function in the female, especially in regard to the proestrous LH surge and estrous cyclicity in mice. Since these adverse effects were manifested under conditions when plasma lithium concentrations were within or around the therapeutic range, our results provide important conceptual information concerning possible adverse effects of lithium on the reproductive function in the human female.

Lithium: short-term and chronic effects on plasma testosterone and luteinizing hormone concentrations in mice.

The effects of short-term and chronic lithium administration on the concentrations of plasma testosterone (T) and luteinizing hormone (LH) were evaluated in C57BL/6 mice, maintained on a fixed photoperiod of LD 14:10 (white lights on at 06:00 h, CST). Lithium chloride was injected intraperitoneally twice daily (at 09:00 and 16:00 h) in groups of adult male mice at a dosage of 2.5 meq/kg for 7 days, and 1.25 meg/kg for 21 days. Circulating levels of T and LH were measured by standard radioimmunoassay (RIA) methods. Plasma T levels showed a significant increase in mice treated with lithium for 7 days as compared to those in saline-injected control animals. However, there was no significant difference in the concentrations of plasma T between chronic (21 days) lithium-treated mice and the matched control. Plasma LH levels remained unchanged following both short-term and chronic lithium treatment.

Lithium-induced changes in the plasma and pituitary levels of luteinizing hormone, follicle stimulating hormone and prolactin in rats.

Adult male Sprague-Dawley rats, maintained under a controlled photoperiod of LD 14:10 (white lights on at 06:00 h, CST), were injected with lithium chloride and changes in the levels of plasma and pituitary homogenates of luteinizing hormone (LH), follicle-stimulating hormone (FSH) and prolactin (PRL) were examined to evaluate the effects of this anti-manic drug on reproductive function. Two groups of rats were injected with lithium chloride intraperitoneally, twice daily at 09:00 and 16:00 h, for 2 and 7 days at a dosage of 2.5 meq/Kg body weight. Plasma and pituitary levels of LH, FSH and PRL were measured by radioimmunoassay. Plasma levels of LH were significantly (P less than 0.05) increased after 2 days of lithium treatment. In contrast, a significant (P less than 0.005) reduction in plasma levels of LH was evident when lithium injections were continued for 7 days. The plasma levels of FSH remained unaffected by lithium treatment by either time period. Lithium administered for 2 days did not bring about any significant alteration in the plasma levels of PRL, although there was a significant (P less than 0.002) reduction in plasma PRL levels after 7 days treatment. The concentrations of pituitary LH, FSH and PRL remained unchanged after 2 and 7 days of lithium treatment.

An ultrastructural comparison of implantation in young and senescent golden hamsters.

Implantation was compared in young (3-5 month) and senescent (14-17 month) golden hamsters at 3, 3 1/2, 4, 4 1/2, 5, and 5 1/2 days of pregnancy (post-ovulation). One hundred twenty-two embryos (89 from 22 young females and 33 from 18 senescent females) located in utero by cutting thick sections (1-3 mum), were re-embedded and thin-sectioned for electron microscopy. Ultrastructurally implantation and embryonic differentiation was delayed approximately 12 hours in the senescent hamster when compared with young hamsters at the same stage of pregnancy. Failure of the blastocyst to implant would not appear to be a major cause of the declining litter size in aged golden hamsters.

The effect of unilateral testicular torsion on the contralateral testicle in prepubertal Chinese hamsters.

Recent studies of experimental testicular torsion in rats, rabbits, and guinea pigs have demonstrated conflicting evidence regarding contralateral testicular damage. Those studies in which cellular damage has been found are postulated to result from an immunological mechanism whereby the blood-testis barrier is disrupted with subsequent autoantibody formation. In this study, the histologic and immunologic effects of testicular torsion on the contralateral testicle were investigated in prepubertal Chinese hamsters. Four study groups were established; (1) Left orchiectomy only, (2) sham surgery (scrotal incision), (3) 720 degrees left testicular torsion with left orchiectomy 24 hours later, (4) 720 degrees torsion of left testicle with detorsion after 24 hours. The initial procedure was performed at 1 month of age with subsequent biopsies of the contralateral testicle at 1 week, 1 month, and 6 months after the initial procedure. Testicular tissue was examined for immunofluorescent activity using fluorescent labeled goat anti-hamster IgG. Positive controls were established by rabbit immunization (rabbit anti-hamster immunoglobulin) which was subsequently combined with fluorescent labeled goat antirabbit IgG. There was no appreciable difference in immunologic activity between control and experimental animals. Representative sections were examined histologically and no tubular damage was demonstrated and active spermatogenesis was noted at 6 months in all groups. We believe that our results support the premise that testicular torsion in the prepubertal period has no effect on the contralateral testicle.

Fertilizing ability of spermatozoa from aged C57BL/6NNia mice.

Spermatozoa from C57BL/6NNia mice (7- and 25-month-old males that produced offspring and 25-month-old males incapable of producing offspring which either mated or did not mate after being paired for 1 month with proven-fertile females) were tested in in-vitro fertilization studies. The 7-month-old males fertilized the largest number of oocytes (80-86%) in vitro and 79% of them subsequently developed into blastocysts in culture. Aged males which failed to mate fertilized the lowest number of oocytes (11-19%) with 48% developing to blastocysts. This group of mice had the lowest number of spermatozoa in the cauda epididymidis (3.2 +/- 0.4 x 10(5)/mg tissue) with fewer motile spermatozoa (22.3 +/- 5.1%) than younger males. The percentage of spermatozoa retaining their acrosome after 3 h in culture was higher in aged males which had not mated when compared to younger males that had mated. After 4 h in culture, however, the number of spermatozoa that had lost their acrosome was almost identical in the two groups. Superovulated mice which were artificially inseminated with spermatozoa from 25-month-old mice that had not mated did not become pregnant. Testosterone concentrations were lowest in aged mice not mating. These concentrations may explain the poor behavioural response of these males, but whether they account for the inability of spermatozoa to fertilize ova in vitro or in vivo after artificial insemination is not known.