Sexual differences in 5'-deiodinase activity in the Harderian gland of Syrian hamsters and the effect of pinealectomy: regulation by androgens.

Sexual differences on thyroxin 5'-deiodinase (5'-D) in the Harderian gland of Syrian hamsters were investigated. We compared the 24-h profile of 5'-D activity in male and female hamsters, observing a clear rhythm in males but not in females. Female values were always significantly higher than male ones. After pinealectomy day/night variations in male 5'-D activity at the time points studied were abolished, results that are in correlation with serum thyroid hormones. We also studied the regulation by androgen of the enzyme activity. Basal 5'-D activity increased in castrated males and levels fell when animals were implanted with testosterone or its product 5 alpha-dihydrotestosterone (DHT). Female 5'-D activity was also inhibited by androgens. As only the addition of DHT in the presence of epitestosterone, an inhibitor of the conversion of testosterone on DHT, in castrated males was able to decrease 5'-D activity to the control animal levels, we suggest a probable direct effect of DHT by itself.

Mg(2+)-dependent and Ca2+, Mg(2+)-dependent ATPase activities in the harderian gland of rodents: age and sex influences.

Three experiments employing male and female Syrian hamsters (aged 1, 2, and 8-10 months), male Sprague-Dawley rats (aged 1, 2, and 10 months) and male C57B1 mice (aged 2, 7, 13, and 29 months) examined the effects of age and sex on Mg(2+)-dependent and Ca2+, Mg(2+)-dependent ATPase activity in the Harderian gland. Significant differences due to age and sex were observed in the hamsters and rats but not with age in mice. Generally, male hamsters had significantly higher Mg(2+)-dependent and Ca2+, Mg(2+)-dependent (exception at one timepoint) ATPase activity than did females. Age-matched male and female rats had similar values of Mg(2+)-dependent ATPase activity, but males had significantly higher Ca2+, Mg(2+)-dependent ATPase activity than females at 2 months of age.

Control of secretory lipid droplets in the harderian gland by testosterone and the photoperiod: comparison of two species of hamsters.

Harderian glands of mammals secrete lipid. They are markedly sexually dimorphic in Syrian (golden) hamsters (Mesocricetus auratus): female glands consist almost entirely of one cell type (type I) with small lipid droplets, whereas glands of males have both type I and type II cells, with large lipid droplets. Siberian (Djungarian) hamsters (Phodopus sungorus) have sexually monomorphic Harderian glands, with both type I and type II cells. We used a morphometric technique to quantify the proportions of small (type 1) and large (type 2) lipid droplets in these two species, in relation to the presence or absence of testosterone and to variations in the photoperiod. In Syrian hamsters, orchidectomy led to a marked increase in the proportion of type 1 lipid droplets in males kept in long (but not short) day photoperiods. In contrast, treatment of females with testosterone led to an increase in type 2 lipid droplets. Short-day photoperiods in both sexes led to an increase in the proportion of type 2 lipid droplets and this was prevented by pinealectomy. In Siberian hamsters, on the other hand, castration or short photoperiods had no effect on Harderian gland morphology in either sex. These results suggest that some property of type 2 lipid droplets is important to Syrian hamsters during the autumn and winter. Syrian hamsters have a dimorphic Harderian gland and testosterone maintains the basic sexual dimorphism during the long days of spring and summer; a pineal-mediated mechanism, perhaps the drop in serum prolactin levels, leads to an increase in type 2 lipid droplets with the short days of autumn and winter.(ABSTRACT TRUNCATED AT 250 WORDS)

Pineal and thyroid functions in newborn seals.

Daily variations of pineal and plasma melatonin and plasma thyroid hormones were measured in harp seals (Phoca groenlandica), grey seals (Halichoerus grypus), and hooded seals (Cystophora cristata), ranging in age from newborn to 14 days. In newborn harp seals the mean mass of the pineal gland was 273 mg (+/- 45 SEM, n = 11), containing 49 ng (median) melatonin. In newborn, 4- and 10-day-old grey seals, the pineal mass was similar, weighing on average 337 mg (+/- 74, n = 6) and containing 90 ng melatonin. Two newborn hooded seal pups had pineals weighing 520 and 1289 mg, with 254 and 7600 ng melatonin, respectively. There were no day-night differences in the pineal contents of melatonin or in the number of pineal beta-adrenergic receptors measured in newborn harp seals, and, in newborn, 4- and 10-day-old grey seals, there were no day-night or age differences in pineal melatonin content. Plasma melatonin levels were 10 times higher in newborn seals than in two 10-day-old grey seals and one 14-day-old harp seal pup. In all seal pups, the levels exhibited a 24-hr rhythmicity, with increasing night- and decreasing daytime concentrations. Plasma levels of thyroxine (T4) and triiodothyronine (T3) were generally higher in newborn seals than in 10- and 14-day-old seals or in adult females. There was no apparent 24-hr rhythmicity, but the thyroid hormone levels generally declined throughout each sampling sequence. High pineal and thyroid activities may play a thermoregulatory role in newborn seals, but the results do not indicate a stimulatory action of melatonin in the peripheral conversion of T4 to T3. It is speculated that the large and active pineal gland, particularly in newborn seals, may be related to aspects of their diving habit.

Melatonin inhibits luteinizing hormone releasing hormone (LHRH) induction of LH release from fetal rat pituitary cells.

The in vitro effect of melatonin on the release of luteinizing hormone (LH) and follicle stimulating hormone (FSH) from fetal rat pituitary cells was investigated. A significant inhibition of LH release induced by 10(-9) M luteinizing hormone releasing hormone (LHRH) was seen when cells were incubated with 10(-9) M melatonin. FSH release was unaffected by either LHRH alone or LHRH in combination with melatonin. In addition, the significant inhibitory effect of melatonin was reduced by pretreatment of the pituitary cells with 10(-10) M melatonin. These findings indicate that melatonin can act directly on the fetal pituitary gland to suppress LHRH-induced release of LH perhaps by a mechanism which eventually involves down-regulation of the melatonin receptors.

Circadian rhythms in reproductive and thyroid hormones in gonadally regressed male hamsters exposed to natural autumn photoperiod and temperature conditions.

Two groups of young adult male Syrian hamsters were kept in a vivarium at 22 degrees C and a light:dark cycle of 14.5:9.5 h (lights on 06.30 h; indoor) or in a naturally decreasing photoperiod and fluctuating ambient temperature conditions (outdoor) from October 1 (day length 11 h 50 min) to November 30 (day length 10 h 12 min). Representative animals from each group were killed at 3-hour intervals with additional time points near the onset of light. Weights of the paired gonads and accessory organs revealed that all of the animals kept outdoors and none of those kept indoors underwent reproductive regression. Significant circadian rhythms were observed in serum luteinizing hormone (LH), follicle-stimulating hormone (FSH), prolactin (PRL), thyrotropin (TSH), thyroxine (T4), triiodothyronine (T3) and testosterone in indoor and outdoor-housed hamsters. The 24-hour acrophase in serum LH, TSH, T4 and T3 occurred between 13.00 and 16.00 h, while that of serum testosterone and PRL occurred between 18.00 and 20.00 h in indoor hamsters. Hormonal variables in which there was a significant alteration in the 24-hour acrophase of outdoor animals relative to that in the indoor animals included pituitary PRL and serum testosterone, PRL, FSH and TSH. Hamsters housed indoors had a significant rhythm in brown adipose tissue type-II 5'-deiodinase activity, but no rhythm was evident in this tissue in outdoor animals. The natural autumnal conditions depressed serum LH and testosterone around the clock, though the depression of serum FSH relative to indoor hamster values was best seen between 09.00 and 21.00 h and that for PRL between 15.00 and 24.00 h.(ABSTRACT TRUNCATED AT 250 WORDS)

Insulin-like growth factor-1 in Syrian hamsters: interactions of photoperiod, gonadal steroids, pinealectomy, and continuous melatonin treatment.

Four experiments in Syrian hamsters examined the role and possible interaction of photoperiod, gonadal steroids, and the pineal on circulating levels of insulin-like growth factor-1 (IGF-1). In the first experiment, female hamsters were exposed to long photoperiod (LP; 14:10 LD) or short photoperiod (SP; 8:16 LD); an additional group of SP-exposed females was pinealectomized (PX). SP induced a significant depression in IGF-1 concentrations which PX partially prevented. In Experiment 2, two groups (control and castrate [CX]) of adult male hamsters were kept in LP, and three groups (intact, CX, and CX+PX) of hamsters were kept in SP for five weeks. The four groups of animals that were CX and/or maintained in SP had approximately the same mean level of IGF-1, and all four groups were significantly (P < 0.001) higher than the LP-control hamsters. In Experiment 3, four groups (intact controls, CX, CX+melatonin pellet [MEL PEL], and MEL PEL only) were kept in LP. Melatonin pellets (1 mg melatonin/24 mg beeswax/every two weeks) were implanted sc twice during the experiment. Castration induced a rise (P < 0.001) in IGF-1 levels, and this was not prevented by MEL PEL. In Experiment 4, testosterone and dihydrotestosterone pellets implanted in LP-exposed CX males prevented the CX-induced rise in IGF-1; testosterone implants also reduced IGF-1 levels in CX males treated with progesterone. In conclusion, SP treatment depresses IGF-1 in female hamsters and raises it in males. These results substantiate previous studies in other models of gonadal steroid deficient animals. They lend further credence to the hypothesis that there is a sexual dimorphism in circulating IGF-1 concentrations in the Syrian hamster that may be at least partially related to the presence of gonadal steroids.

Pineal acid phosphatase activity in Syrian hamsters: sex differences and effects of castration and androgen replacement therapy.

Pineal acid phosphatase (ACP) activity was examined in seven experiments involving young intact or castrated male and female Syrian hamsters. Nine-week old female hamsters had a 3-fold (p < 0.001) higher ACP activity in their pineal glands than did males. After three weeks of castration, a significant increase (p < 0.001) in ACP activity occurred in castrated male pineal glands. In males, pellets with 5 alpha-dihydrotestosterone (p < 0.01) but not testosterone or progesterone suppressed pineal ACP activity. In females, no changes in pineal ACP activity were noted due to the estrous cycle. Pineal ACP activity was not affected by testosterone, dihydrotestosterone or androstenedione pellets in intact females or by testosterone pellets in ovariectomized animals.

Development and androgen regulation of the secretory cell types of the Syrian hamster (Mesocricetus auratus) Harderian gland.

The secretory cell types of the hamster Harderian glands were studied in both male and female Syrian hamsters. As previously demonstrated, female hamsters showed a single secretory cell type (type I), while male hamsters displayed two secretory cell types (type I and type II). Type-II cells were observed after the first month of age correlating with the increase in testosterone levels. The administration of testosterone to adult female hamsters resulted in a marked increase in the percentage of type-II cells without a significant increase in the number of mitotic figures. Very low levels of serum testosterone were able to maintain the percentage of type-II cells. Castration of male hamsters produced a decrease in the percentage of type-II cells. This drop correlated with the reduction in serum testosterone levels. The chronic administration of a luteinizing hormone-releasing hormone agonist to male Syrian hamsters induced a significant reduction in both serum luteinizing hormone and testosterone. However, the percentage of type-II cells was similar to that of control hamsters suggesting that very low levels of circulating testosterone are able to maintain the percentage of type-II cells. In a final experiment male Syrian hamsters were treated with the antiandrogen cyproterone acetate. No changes were observed in the percentage of type-II cells, whereas serum luteinizing hormone and testosterone levels were significantly modified. We concluded that (1) type-II cells differentiate from type-I cells; (2) gonadal androgens are the major factor controlling this differentiation; and (3) the disappearance of type-II cells after androgen deprivation occurs through holocrine and apocrine mechanisms.(ABSTRACT TRUNCATED AT 250 WORDS)

Gender-associated differences in the development of 5-aminolevulinate synthase gene expression in the harderian gland of Syrian hamsters.

The mRNA levels for aminolevulinate synthase (ALV-S), the rate-limiting enzyme in porphyrin synthesis, were studied in male and female Syrian hamsters during postnatal development. Sex-associated differences in the expression of ALV-S gene were evident at the end of the third week of postnatal development. Serum levels of luteinizing hormone (LH), testosterone, cortisol, thyroid hormones and insulin-like growth factor were also studied in order to correlate their concentrations with the mRNA levels for ALV-S. Among these hormones, serum LH levels showed a positive correlation with the ALV-S mRNA levels. However, the expected negative correlation with testosterone levels was not clearly observed. Thus, in order to test the effects of testosterone on ALV-S gene expression, 11-day-old male and female Syrian hamsters and adult female hamsters were injected with 50 micrograms of testosterone for 4 days. Testosterone administration decreased the levels of ALV-S mRNA in the adult females but did not influence those of young females. The possible explanation for the insensitivity to testosterone during these postnatal stages might involve the maturational state of androgen receptors in the Harderian glands.

Chronic N-methyl-D-aspartate administration prevents melatonin-associated changes in cell differentiation in the harderian glands of male hamsters.

The daily administration of 25 micrograms of melatonin for 10 weeks resulted in an increase in the percentage of Type II cells in the Harderian glands of male Syrian hamsters. Harderian glands of melatonin injected animals consisted of 65-70% Type II cells while control animals which were injected with saline had 40% Type II secretory cells. The daily administration of 3 mg of the glutamate receptor agonist N-methyl-D-aspartate (NMDA) prevented the effects of melatonin on cell differentiation but was without effect when administered to saline treated hamsters alone. Both the relative number of mitoses and the number of total cells, estimated by counting the nuclei, was not affected. Thus, a conversion from Type I to Type II cells seems possible. The effects of melatonin and NMDA administration were independent of the serum levels of testosterone, luteinizing hormone and thyroxine, hormones which have been implicated in Type II cell differentiation. However, prolactin levels, which were affected by melatonin and NMDA administration, might be involved in the differentiation of Harderian gland secretory cells.

Type II iodothyronine 5'-deiodinase in mouse brown adipose tissue is stimulated by a single injection of isoproterenol in an oil/water emulsion.

A single subcutaneous morning injection of the beta-adrenergic agonist isoproterenol (ISO) (10 mg/kg body weight) in an oil/water emulsion (70/30; v/v) caused a marked increase in the activity of the enzyme type II iodothyronine 5'-deiodinase (5'-D II) in the interscapular brown adipose tissue of BDF-1 mice. After a delay of 4 hours, the 5'-D II activity began to rise in an almost linear fashion and was increased 3-fold after 8 hours, when compared to the control values. The results indicate that this method of ISO administration may be a valid tool for artificial stimulation of brown adipose tissue in animals by beta-adrenergic agonists.

Testosterone sensitive dihydropyridine binding in the Harderian gland of the male hamster.

It is well known that in different tissues, dihydropyridines bind at nanomolar concentrations to a receptor and block voltage-operated Ca2+ channels. In studies reported here, Harderian gland tissue homogenates from intact male hamsters exhibited significant dihydropyridine binding (Bmax = 1700 fmoles/mg protein) of high affinity (Kd = 1.1 nM). Tissue homogenates from female animals exhibited a similar Kd value (1.35 nM) but receptor density per mg protein was significantly reduced (Bmax = 270 fmoles). Dihydropyridine binding of Harderian gland tissue homogenates from castrated males was reduced greater than 80% (Bmax = 225 fmoles/mg protein). Treatment of castrated males with subcutaneous testosterone pellets resulted in significant restoration of dihydropyridine binding activity (approximately 80%, Bmax = 1630 fmoles/mg protein) with a comparable binding constant (Kd = 1.50 nM) as observed for noncastrated, control animals. Addition of testosterone (ex vivo) to homogenates from castrated hamsters did not restore dihydropyridine binding to control levels. These data indicate: (a) the Harderian gland from male hamsters exhibits significant dihydropyridine binding; (b) ligand binding is abolished following castration; and (c) significant restoration of dihydropyridine binding occurs following in vivo testosterone treatment. The dependence of dihydropyridine binding restoration upon in vivo steroid hormone administration suggests probable involvement of the steroid at the transcriptional level although non-genomic mechanisms such as the binding of testosterone to a receptor resident in the plasma membrane and subsequent activation of Ca2+ channels can not be ruled out.

Porphyrin metabolism in the harderian glands of Syrian hamsters: in vivo regulation by testicular hormones, lighting conditions, pineal gland, and pituitary hormones.

The porphyrin concentration in the harderian glands of male hamsters subjected to several endocrine manipulations was studied. Prolonged bilateral gonadectomy resulted in a marked increase in harderian porphyrin concentration. This change was not prevented by either pinealectomy or by constant white light exposure. Castrated hamsters exposed to constant red light showed higher porphyrin concentrations than castrated hamsters kept under white light. Among several hormones studied, serum luteinizing hormone and thyroid-stimulating hormone levels were unexpectedly higher in the constant red light exposed group than in the other groups. In order to test whether luteinizing hormone was involved in the postcastrational rise in harderian porphyrins, we administered a potent luteinizing hormone-releasing hormone (LHRH) agonist. The chronic administration of the LHRH agonist resulted in a decrease in serum luteinizing hormone (because it desensitized the LHRH receptors on the gonadotropes) and, consequently, in serum testosterone levels. However, no rise in harderian porphyrin was observed. It is concluded that the absence of testicular hormones might not be the triggering factor involved in harderian porphyrogenesis.

Hypophysectomy prevents the castration-induced increase in porphyrin concentrations in the harderian glands of the male golden hamster: a possible role for prolactin.

The Harderian glands of golden hamsters contain high concentrations of porphyrin pigments, with female hamsters having considerably higher porphyrin concentrations than males. Castration of male hamsters leads to a rapid increase in porphyrin concentrations; testosterone treatment of females has the opposite effect, suggesting a central role for androgens in inhibiting the realization of high porphyrin concentrations by this organ. Previous studies in our laboratories have shown, however, that administration of a dopamine agonist to castrated hamsters prevents the normal increase in Harderian porphyrins from occurring. This suggests that prolactin is necessary for low androgen levels to lead to maximal increases in Harderian porphyrin concentrations. The present study tested the hypothesis that prolactin is involved in the control of Harderian porphyrin levels in the golden hamster. Although hypophysectomy of male hamsters reduced serum testosterone to levels in castrated hamsters, the resultant increase in Harderian porphyrin concentrations was much less than that seen after a similar period of castration. Furthermore, combining the two procedures (castration and hypophysectomy) also led to a blunted increase in Harderian porphyrin, suggesting that a pituitary hormone is necessary for low testosterone levels to lead to increased porphyrins. Evidence that this pituitary hormone is prolactin comes from the observations that eliminating all pituitary hormones except prolactin, by severing the connection of the pituitary with the hypothalamus or transplanting the pituitary to a distant site (beneath the kidney capsule) led to greatly augmented Harderian porphyrin levels, in intact or castrated male hamsters.(ABSTRACT TRUNCATED AT 250 WORDS)

N-methyl-D-aspartate does not prevent effects of melatonin on the reproductive and thyroid axes of male Syrian hamsters.

The reproductive and thyroid status of male Syrian hamsters maintained on long days (14 h light, 10 h darkness) were assessed after 10 weeks of daily injections of pharmacological doses of melatonin (25 micrograms s.c.) and/or N-methyl-DL-aspartic acid (NMDA, 0.025-6 mg i.p.), a compound with receptor sites in the central nervous system which are known to affect reproduction. Melatonin given during the late light phase decreased reproductive organ weights and levels of serum and pituitary prolactin and serum thyroxine (T4); these results are similar to published reports on the effects of chronic short photoperiod treatment of this species. Reproductive organ weights, T4 levels and values for prolactin did not differ significantly between groups receiving only melatonin and those receiving NMDA in addition to melatonin; likewise these variables did not differ significantly between groups receiving only either NMDA or saline. NMDA alone and in combination with melatonin increased serum tri-iodothyronine (T3). The brown adipose tissue enzyme T4 5'-deiodinase demonstrated an increased activity in the presence of NMDA, with the lowest dosage eliciting the most significant effect. Previous studies have demonstrated that NMDA reverses the reproductive effects of short photoperiod. The results of this study show that NMDA is incapable of preventing the inhibitory reproductive effects of exogenously administered melatonin. These observations are consistent with the proposal for a site of action for NMDA on neural regions more proximal than those altered by melatonin; alternatively, NMDA may interfere with neurotransmitter actions in the pathway controlling melatonin production.

Serum luteinizing hormone, prolactin, and thyrotropin and their pituitary subunit mRNA levels during proestrus in the Syrian hamster.

mRNA levels for alpha, luteinizing hormone beta (LH beta), and prolactin (Prl) were examined during the hamster estrous cycle, with sampling most frequent (1-hour intervals) on the afternoon of proestrus. These transcripts encode the peptide subunits for the pituitary hormones LH and Prl which are necessary for reproductive function. Serum hormone levels of LH and Prl, analyzed by 24-hour periodic regression, exhibited a 24-hour periodicity on proestrus characterized by a large surge peaking at about 18.00 h. Combining the data for non-proestrous days of the cycle disclosed a rhythm with similar timing for LH and Prl. Thyroid-stimulating hormone (TSH) and TSH beta RNA profiles during hamster proestrus are reported for the first time. Serum TSH exhibited a pronounced peak coincident with that of the other hormones on proestrus. Because of variations at other times on the day of proestrus, however, a 24-hour periodicity was not manifested by regressional analysis. Combined non-proestrous serum TSH data also revealed no consistently timed regressional 24-hour periodicity. During proestrus, pituitary mRNA values for alpha, LH beta, and Prl simultaneously exhibited a rise from the lowest to the highest of all proestrous values in the 3-5 h prior to the time of the pre-ovulatory peak of circulating hormone concentrations. RNA for TSH beta exhibited an earlier, broader peak on proestrus. Periodic regression indicated a significant 24-hour rhythm for alpha mRNA in data pooled from non-proestrous days (acrophase 05.00 h) and for TSH beta mRNA on proestrus (acrophase 04.54 h).(ABSTRACT TRUNCATED AT 250 WORDS)

Endocrine and metabolic effects of life-long food restriction in rats.

The effects of chronic (40%) food restriction from 6 weeks of age were studied in 28-month-old male Fischer-344 rats; the results were compared with ad libitum-fed, old and young male rats at 28 and 3 months of age, respectively. Pituitary luteinizing hormone was similar in all old rats and was significantly lower than in young rats. In old ad libitum-fed, but not in food-restricted rats, serum levels of LH, testosterone and T4 were significantly lower than in young rats. Serum levels of T3 did not differ between young and old rats. Type-II 5'-deiodinase activity in brown adipose tissue was similar in both groups of old animals and was significantly depressed as compared with that in young rats. Serum levels of triglycerides were significantly depressed in food-restricted rats, but were significantly increased in ad libitum-fed rats as compared with young rats. Both groups of old rats had significantly elevated serum levels of cholesterol over that in young rats, but the level was significantly lower in food-restricted as compared to ad libitum-fed animals. The results are consistent with the notion that life-long food restriction tends to preserve the activity of many metabolic functions.

Food restriction retards aging of the pineal gland.

The effects of chronic (40%) food restriction from 6 weeks of age were studied in aging male Fisher 344 rats. When compared with 3-month-old, ad libitum fed rats, pineal N-acetyltransferase (NAT) activity had declined to less than 30% and pineal and serum levels of melatonin to 40% after 28 months when feeding had been ad libitum. Food restriction significantly retarded this development (P less than 0.05) giving NAT and melatonin levels which were twice as high as in the ad libitum fed group. Nighttime levels of pineal serotonin (5-HT) were similar in food-restricted and ad libitum fed old rats but were nearly twice as high (P less than 0.05) as in young rats. There was also a tendency for increased production of 5-hydroxyindoleacetic acid (5-HIAA) in the pineal gland with higher levels of 5-HT. It is concluded that aging in the rat (Fisher 344) is accompanied by a reduction of pineal NAT activity, thereby reducing the production of melatonin and causing a buildup of 5-HT in the pineal gland. It is furthermore proposed that food restriction, which markedly increases the life span and reduces age-related physiological deterioration and diseases in many animals, may mediate some of its effects through a sustained pineal activity in old age.