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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.

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.

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 (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.

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.

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.

The effect of 24-hr exposure to cold (5 degrees C) was studied in male Syrian hamsters adapted to short days (LD 8:16). Both pineal N-acetyltransferase (NAT) activity and pineal and serum concentrations of melatonin showed a clear, diel rhythm with a moderate but significant increase late in the dark period. The nighttime peak levels of NAT activity and pineal and serum melatonin were significantly higher in the animals that had been exposed to cold than in those remaining in warm (22 degrees C) conditions. The activity of type-II 5'-deiodinase (5'D) in the brown adipose tissue (BAT) was not different after 24 hr of cold exposure between cold- and warm-exposed hamsters, and the enzyme did not show any diel rhythmicity. It has been speculated that some effects of cold exposure may be simulated by melatonin treatment; the present data further support this notion. The apparent lack of response in BAT 5'D activity remains enigmatic and needs further investigation.

Circadian rhythms in acid phosphatase (ACP), hexosaminidase (HEX) and beta-glucuronidase (RON) activity were studied in the pineal glands of adult male Syrian hamsters exposed to control (20 +/- 2 degrees C and 14:10 LD) conditions or to naturally decreasing autumn photoperiod and temperature conditions (outside) for 8 weeks. Testes and testosterone levels (p less than 0.001 in both instances) were severely depressed in animals exposed to natural environmental conditions illustrating that the treatment period was of sufficient length to produce pineal-mediated gonadal atrophy. Significant rhythms were found in all enzymes in the pineal glands of control and outside animals with the exception of HEX activity in the control animals. Significant acrophase differences in outside vs. control animals were noted in ACP (7.9 hr earlier, p less than 0.001) and RON (9.8 hr later, p less than 0.001). A significant drop in RON and HEX activity (p less than 0.01 in both instances) was noted in association with the acute lights exposure in the morning to which control animals were exposed. The around-the-clock mean value of each enzyme was significantly lower in outside vs. control hamsters. These results demonstrate that environmental changes which provoke the pineal-mediated depression in gonadal activity also alter the activity of and shift the circadian rhythmicity of lysosomal enzymes in the pineal itself.

1. Pineal N-acetyltransferase (NAT) activity in long-day white-footed mice (Peromyscus leucopus) exposed to cold soon after onset of darkness was depressed relative to controls, whereas mice exposed to cold later at night had slightly elevated enzyme activity. 2. NAT activity in short-day mice exposed to cold soon after lights off did not differ from controls. Pineal melatonin in these mice, however, did not rise, as it did in controls. 3. These results suggest that acute cold exposure may modulate NAT activity, which is controlled primarily by the L:D cycle.

A circadian rhythm in acid phosphatase and hexosaminidase was found in adult male hamsters exposed to a long photoperiod (14:10 h light/dark [LD]; lights on 06.00 h) and killed at 08.00, 14.00, 20.00, 02.00, 04.00, 05.50 and 0.615 h. Hexosaminidase and beta-glucuronidase activity at 02.00, 04.00 and 05.50 h (values pooled for these times before lights on) were significantly elevated compared to enzyme activity at 06.15 and 08.00 h (pooled values after lights on), suggesting a fall in activity associated with lights on. Hypogonadism was induced in female Syrian hamsters by exposure to a short photoperiod (10:14 h LD) until a majority of them were vaginally acyclic. Pineal lysosomal enzyme activities (acid phosphatase, beta-glucuronidase, hexosaminidase, alpha-arabinosidase and beta-galactosidase) were significantly elevated in short photoperiod-exposed animals compared to animals in 14:10 LD, when measured near the middle of the light phase. In the third experiment, castrated animals were used to determine if lowered androgen levels might also affect pineal lysosomal enzyme activity. The results indicated that light phase beta-glucuronidase, hexosaminidase and beta-glucosidase activities were lower in castrated males compared to their intact controls. In summary, these results demonstrate that (1) lysosomal enzyme activity is present in the Syrian hamster pineal, (2) changes can be observed which suggest involvement of this activity in pineal function and, (3) a circadian rhythm in enzyme activity is present with peak activity occurring during the night. In the short photoperiod and castration experiments, the changes in lysosomal enzyme activity could reflect either a hormonal manipulation or a change in circadian regulation of enzyme activity.(ABSTRACT TRUNCATED AT 250 WORDS)

Previous studies show that the suppression of gonadal function in blind-anosmic rats is dependent on the pineal gland. The present results demonstrate that in young female rats both the pineal gland and dual sensory deprivation have additional independent antigonadal effects.

Melatonin synthesis in the pineal gland, which is primarily regulated by the environmental lighting regime, can also be influenced by other factors that elicit modifications in sympathetic tone. The objectives of this study were to determine if forced swimming alters the normal pattern of melatonin production in the pineal gland of the Richardson's ground squirrel (Spermophilus richardsonii). In early June, the squirrels were forced to swim for 10 min during the photophase or during the scotophase. In mid-July squirrels swam only during the scotophase. Animals were sacrificed 15, 30, or 60 min after the onset of swimming. Activities of pineal N-acetyltransferase (NAT) and hydroxyindole-O-methyltransferase (HIOMT) were assessed by radioenzyme assay, and pineal melatonin content was measured by radioimmunoassay. Daytime swimming elicited no major changes in enzyme activity or pineal melatonin. In June, swimming at night prevented the normal rises in NAT activity and pineal melatonin seen in nonswimming controls. In contrast, the pineals of squirrels that were tested 6 weeks later in mid-July did not appear to be as sensitive to nighttime swimming, as there were only minor differences in both NAT activity and melatonin content compared to controls. These results demonstrate that forced nighttime swimming, unlike several other aversive stimuli, can evoke changes in the normal pattern of pineal melatonin production in this species. Furthermore, the pineal's response to such stimuli may not be stable over the course of the active season.

Young adult male Syrian hamsters maintained in an LD 14:10 photo-period (lights out at 8 p.m.) were given a single injection of 25 micrograms melatonin or ethanolic-saline at 4 p.m. Animals from each group were sacrificed at 5 p.m., 4 a.m., and 10 a.m. following the injections. Pituitary and plasma LH and prolactin as well as pineal melatonin levels were determined in all animals by RIA. Pineal melatonin levels were significantly (p greater than 0.025) reduced at 10 a.m. in the melatonin treated group. Melatonin administration also suppressed pituitary LH levels at 4 a.m. and plasma LH levels at 10 a.m. Plasma prolactin concentrations at 4 a.m. and pituitary prolactin levels at both 5 p.m. and 10 a.m. were significantly elevated in melatonin treated hamsters compared with those given ethanolic-saline. These results indicate that exogenous melatonin has immediate effects on several hormones important in the regulation of reproductive events in this species. This acute injection model may therefore aid in the elucidation of melatonin's mechanism of action in the Syrian hamster.

Pineal N-acetyltransferase (NAT) activity and radioimmunoassayable levels of melatonin were compared in 2-month-old (young), 12-month-old (middle-aged), and 29-month-old (old) female rats killed at 1600 h (during the light) and at 2300 h (4 h after darkness onset) and 0100 h (6 h after darkness onset). During the light period, NAT levels were equivalent in pineals from each age group. With the onset of darkness NAT levels rose sharply and were again equivalent in all groups at 2300 h. At 0100 h pineal NAT values in the old rats were lower than in the other two groups. Melatonin values were low in pineal glands of all animals killed at 1600 h in light. By 4 h after darkness onset pineal melatonin content in the young rats had increased 12-fold compared to a 6-fold rise in the old animals. Melatonin levels in the middle aged rats were intermediate between the other two groups. Similar relationships were observed in the rats killed at 0100 h. By this time the young rats had melatonin levels 17 times higher than during the day while the increase in the old rats was only 7-fold; 12-month-old rats again had intermediate levels. The finding show a marked reduction in pineal melatonin with aging in female rats.

Karyometric investigations of thyroid follicular cells were performed in four groups of adult male gerbils [I. sham-operated (Sham PX) + short photoperiod (SPP)-exposed; II. Sham PX + long photoperiod (LPP)-exposed; III. pinealectomized (PX) + SPP-exposed; IV. PX + LPP-exposed]. The mean nuclear volume (MNV) of thyroid follicular cells was significantly higher in both PX groups (III and IV) when compared to Sham PX controls (I and II). In Sham PX gerbils the maintenance of animals in SPP (10 h light: 14 h darkness) for 13 weeks resulted in significant decrease of MNV of thyroid follicular cells in comparison to that in Sham PX animals kept in LPP (14 h light: 10 h darkness). Lighting conditions did not significantly affect the MNV of thyroid follicular cells in PX gerbils. These data suggest that the pineal gland itself and not light deprivation is the most important factor depressing the activity of the thyroid in male gerbils. Lighting conditions seem to affect the function of thyroid follicular cells inasmuch as they activate or block the action of the pineal gland.

Sensory deprivation produced by removing the eyes and olfactory bulbs in male rats allowed pinealectomy to markedly augment the post-adrenalectomy elevation of ACTH levels. Pineal removal or sensory deprivation separately did not have this effect. Thus, intact sensory input and an intact pineal gland are independently capable of restricting the post-adrenalectomy rise in ACTH levels.

It has recently been shown that a 1.5-ml subcutaneous saline injection into the dorsal aspect of the hind limb induces a dramatic and rapid fall in N-acetyltransferase activity and melatonin content of the rat pineal gland at night. Since many studies have shown the opposite response to stress during the day, the first experiment was undertaken to test whether the timing of the saline injection at night influences the response of the pineal gland. In the present studies, rats were kept under light:dark (LD) cycles of 14:10 with lights out daily at 2000 h. Groups of rats were then given a saline injection at one of the following times: 2315, 0015, 0115, 0215, or 0315. Early in the dark phase (2315 and 0015) the saline injection depressed both the N-acetyltransferase (NAT) activity and the melatonin content of the pineal. As the animals were treated later in the dark period, the response became more blunted and, finally, disappeared. In the second experiment, animals that were kept in light during the usual dark period showed no pineal response when subjected to a hind leg injection of saline at either 2315 or 0315. Additionally, no response was seen in the two pineal parameters when rats had darkness onset delayed by 4 h (to 2400) and were then treated with saline at 0410. The results of these studies indicate that the pineal response to saline injection is time dependent. Also, if the nighttime rise in melatonin is suppressed by light exposure, a saline injection has no further inhibitory effect on pineal NAT activity or melatonin levels.

Previous observations have suggested a role for the pineal gland in regulation of blood pressure. To investigate this possibility, pinealectomy was performed in prepubertal (43-day-old) and postpubertal (55-day-old) male rats. Systolic blood pressure (BP) was measured by the tail cuff method in conscious pinealectomized (Px) and sham pinealectomized (ShPx) rats at various times. In the 3 weeks following surgery, the BP increase after Px was twice that after ShPx (p less than 0.05). During administration of 1% saline as the sole fluid source from day 96-123, hypertension (BP greater than 150 mmHg) was observed in those animals Px at day 43 (p less than 0.05 vs ShPx) but not in those Px on day 55. Moreover, saline consumption was increased in the hypertensive group compared to ShPx controls. There was no significant difference in mean body weight or heart rate between Px or ShPx groups. These data suggest that pinealectomy in the prepubertal period may result in a tendency to hypertension, measurable in conscious rats, which is unmasked by saline administration.

Blinding adult male golden hamsters led to involution of the testes and accessory sex organs (seminal vesicles and coagulating glands) and to a regression in pituitary prolactin levels within 8 weeks. The subcutaneous implantation of either melatonin or 6-hydroxymelatonin (1 mg/wk in beeswax) prevented the atrophy of the reproductive organs and the decrease in the stores of pituitary prolactin. Two other indoles, N-acetylserotonin and 5-hydroxytryptophol, failed to counteract the reproductive effects of blinding. Both melatonin and 6-hydroxymelatonin significantly elevated plasma LH titers.

The pineal gland of four-month-old gerbils contains numerous calcareous deposits (corpora arenacea or concretions). Also, the tissue appears vacuolated due to the presence of a number of large rounded spaces. The deposits are usually associated with or lying within these "vacuoles". By eight months of age, the number of both the corpora arenacea and the "vacuoles" are increased over those present in the pineals of four-month-old gerbils. Bilateral superior cervical ganglionectomy at one month of age prevents the formation of both the concretions and "vacuoles". The cells in the sympathetically denervated pineal glands are smaller than those in pineal glands of animals that possess intact superior cervical ganglia.

Maintaining adult male golden hamsters in short daily photoperiods (1hr of light and 23hr of darkness daily; LD 1.23) for 12 weeks caused the tests and accessory sex organs to atrophy and also led to significant depressions in pituitary LH and prolactin levels. If hamsters that were kept in LD 1.23 CYCLES RECEIVED WEEKLy subcutanceous implants of either a melanioninbeeswax or a 5-methoxtrypotophol-beeswax pellet (1 mgindole in 24 mg beeswax) the testes and accessory sex organs failed to involute and pituitary LH levels did not drop. Both treatments also retarded the depression in hypophyseal prolactin retarded the depression in hypophyseal prolactin levels. Treatment (twice daily on weekdays and once daily on weekends) with 1.5 mug injections of synthetic LRH (in 0.2 ml 8% gelatin) did not prevent gonadal or accessory organ atrophy while it further depressed hypophyseal LRH treatment, plasma LH titers were significantly elevated. The reproductive organs of hamsters that were moved from short (LD1;I 23) to long (LD 14;10) daily photoperiods regenerated within 8 weeks. This light-induced restoration of the gonads was not prevented or retarded by the weekly implantation of either melantonin-beeswax or 5-methoxytrptophol-beeswax pellets. The results suggest that in the male golden hamster neither melatonin nor 5-methoxtryptophol accounts for the antigonadotrophic activity of the pineal gland.

Estrogen-progesterone (EP) treated adult male rats were injected intravenously (iv) with 0.1, 1 or 10 mug arginine vasotocin (AVT) or Ringers lactate solution. A significant dose-related rise in plasma prolactin was evident 10 min after injection with AVT. In a second experiment, sham-operated or pinealectomized EP-treated male rats were injected iv with 0.1 or 1 mug AVT or diluent. Plasma prolactin was significantly elevated in both sham-operated and pinealectomized groups at both 10 and 20 min post-injection of 1 mug AVT. These results indicate that AVT has prolactin-releasing activity and that this activity is not dependent upon the presence of an intact pineal gland.

Harderian gland (HG) weight and lysosomal enzyme activity were evaluated after 21-day-old female rats were singly caged in a long (LP; 14:10 LD) or short (SP; 8:16 LD) photoperiod and fed on one of two dietary regimens (fed ad libitum or 50% underfed) for 50 days; an additional fed and an underfed group of animals in LP were injected every afternoon with 100 micrograms melatonin. Absolute HG weights were significantly lower in all underfed groups compared to their respective fed controls or to the LP fed control group. Absolute HG weights of underfed rats in SP were significantly lower than the underfed rats in LP. Relative HG weights (mg/100 g body wt) were significantly higher in the underfed saline or melatonin-treated groups compared to their respective fed controls; however, HG of the underfed SP group were not different from SP-fed controls. No significant differences in HG acid phosphatase, hexosaminidase, and beta-glucuronidase activities were observed in any of the treatment groups maintained in LP. Acid phosphatase, hexosaminidase, and beta-glucuronidase activities were significantly elevated in HG of underfed animals maintained in SP compared to their respective fed controls or to the LP-underfed group. Both the underfed control and the underfed-melatonin treated groups had lower pineal protein values than their respective fed groups; underfed animals in 8:16 LD had similar pineal protein values compared to those of the fed control group in SP. Significant effects of photoperiod and underfeeding with no interaction between these variables were observed on pineal acid phosphatase. The fed group maintained in 8:16 LD had significantly higher acid phosphatase activity than the fed group kept in 14:10 LD. In conclusion, underfeeding resulted in severely reduced body weights and absolute Harderian gland weights. Increased activity in certain lysosomal enzymes occurred in both the pineal and Harderian gland and in some instances this was dependent upon the light cycle and dietary regimen to which the animals were exposed.

The thiol reagent cysteamine was administered to adult male rats with the aim of investigating its effect on different neural and pineal components. As expected, immunoreactive somatostatin decreased in the median eminence (ME) (p less than 0.05) and gastric antrum (p less than 0.05) after cysteamine; however, no significant change was observed in the pineal IRS content after drug treatment. A decrease in norepinephrine was observed in the ME (p less than 0.001), hypothalamus (p less than 0.001) and pineal gland (p less than 0.05), together with a rise in ME (p less than 0.005) and hypothalamic dopamine (p less than 0.005) content; these results are consistent with a dopamine-beta-hydroxylase inhibiting effect of cysteamine. No effect was observed on hypothalamic serotonin and 5-hydroxyindole-acetic acid content. Pineal N-acetyltransferase (NAT) activity was significantly higher (p less than 0.05) after cysteamine than after saline, but no statistically significant effect was observed on pineal melatonin content. The mechanism involved in the NAT rise is presumably not related to the known stimulatory effect of norepinephrine, which fell after cysteamine. It is suggested that cysteamine may act at an intracellular level, inhibiting NAT degradation, an effect demonstrated in vitro and thought to be related to a thiol:disulfide exchange mechanism.

Twelve weeks after bilateral superior cervical ganglionectomy, the number of pineal concretions in adult male gerbils was drastically decreased irrespective of photoperiodic treatment. These findings present further evidence that pineal concretions may be related to the metabolic activity of the gland. It is suggested that the pineal concretions of the gerbil are associated, in a yet unknown way, to the metabolism of the gland and that an active pineal is essential to maintain the large number of concretions seen in the normal state.

A hind-leg subcutaneous saline injection into rats at night elicits a decrease in N-acetyltransferase (NAT) activity and melatonin content of the pineal gland. The decrement in pineal melatonin production after saline injection is prevented by adrenalectomy. The present studies were undertaken to determine what factor(s) from the adrenal gland cause(s) the drop in pineal melatonin production after saline injection at night. In the first study, groups of intact and adrenal-demedullated male rats were given a saline injection at 23.10 h (3 h, 10 min after lights off) and their pineals were collected 15 or 30 min later. Pineal NAT activity was depressed in both intact and adrenal-demedullated rats at 15 min postinjection as compared to their respective control animals. Pineal melatonin levels exhibited a drop in intact animals at 15 min and in adrenal-demedullated rats at 30 min. In a second study, hypophysectomy was found to prevent the drop in nocturnal pineal NAT activity and melatonin levels normally associated with a hind leg injection of saline. Finally, in a third experiment, groups of hypophysectomized rats were injected i.p. with corticosterone at 23.10 h and killed 10, 25 or 40 min postinjection. Corticosterone injection in hypophysectomized rats produced a response similar to that caused by saline injection in intact animals: NAT activity was depressed at 10 min and melatonin content was lowered at 25 min. These results suggest that the adrenal-mediated depression in melatonin synthesis after saline injection at night in rats may be elicited by an adrenal cortical hormone (corticosterone) and apparently does not involve the release of factors from the adrenal medulla.

To determine if exogenously administered alpha-melanocyte stimulating hormone (alpha-MSH) affects nighttime pineal N-acetyltransferase activity, pineal levels of 5-hydroxytryptophan, serotonin and melatonin, and plasma prolactin levels, adult male hamsters were injected at 1900 hr (lights out 2000-0600 hr) with two doses of the peptide and killed at 0300 hr. The low dose of alpha-MSH (200 ng) produced a significant fall in pineal serotonin, pineal NAT activity and plasma prolactin values. The high dose of the peptide (20 micrograms) increased circulating prolactin titers and pineal serotonin levels and caused a concomitant decrease in pineal melatonin levels.

1. The pineal melatonin content of photoperiod responsive and photoperiod non-responsive male deer mice was assessed. 2. Pineal melatonin content did not differ between short-day mice with regressed testes or those that maintained their reproductive systems. 3. These data suggest that differential phasing of melatonin release or post-pineal mechanisms are responsible for the differential responsiveness to day length among deer mice.

Light deprivation by blinding in female hamsters was followed by a regression of the reproductive organs, an elevation of pituitary LH concentration and a depression of pituitary prolactin levels. Pinealectomy negated almost completely the effects of light deprivation on the neuroendocrine-reproductive axis. Weekly subcutaneous implants of a melatonin:beeswax pellet completely prevented the pineal gland from inhibiting reproductive physiology in blinded hamsters. The findings suggest that melatonin is not pineal antigonadotrophic factor in female golden hamsters. Melatonin implanted hamsters also had higher than normal levels of plasma prolactin.

In light of recent studies demonstrating stress-induced changes in pineal indoleamine metabolism, we tested the effect of acute cold stress on pineal biosynthetic function. Adult male rats were subjected to 30, 60, or 120 min of cold exposure (Ta = 2 degrees C) during either the light or dark phase of the daily photoperiodic cycle. Controls were kept at room temperature (22 +/- 2 degrees C). Animals were killed by decapitation and pineals were analyzed by radioimmunoassay for melatonin content and by radioenzymeassay for the activity of N-acetyltransferase (NAT). Cold exposure during the day elicited no significant changes in pineal indoleamine metabolism. Exposure to cold for 1 hr during the second hour after lights off slightly increased pineal melatonin content, without a concomitant change in NAT activity. Rats exposed to 2 hr of cold beginning 2 hr after lights off, however, displayed a 50% reduction in NAT activity, whereas pineal melatonin content remained unchanged. The paradoxical response of pineal NAT activity and melatonin content are not uncommon when rats are exposed to adverse stimuli.

Recently, it was shown that a 1.5-ml subcutaneous saline injection depressed N-acetyltransferase (NAT) activity and melatonin content in the rat pineal gland at night. The present studies were undertaken to determine if another perturbation, swimming, could duplicate this response. Rats swam at 23.10 h (lights out at 20.00 h) for 10 min and were killed 15 and 30 min after the unset of swimming. Pineal NAT activity was found to be unaffected while melatonin content was depressed dramatically. Hydroxyindole-O-methyltransferase (HIOMT) activity as well as the content of serotonin (5HT), 5-hydroxytryptophan (5HTP) and 5-hydroxyindoleacetic acid (5HIAA) were not changed by this treatment. In a second study, pineal melatonin again was depressed without a concomitant drop in NAT activity. Mean serum melatonin at 15 min after onset of swimming was increased although the rise was not statistically significant. In the final study, it was found that NAT activity was slightly increased in intact rats and unchanged in adrenalectomized rats at 7 min after swimming onset. At 15 min both intact and adrenalectomized animals had NAT activity values similar to those of controls. Pineal melatonin content in intact and adrenalectomized rats plummeted to 50% of control values at 7 min and fell further to 25% at 15 min. While the rate of melatonin synthesis was not directly measured, lack of change in the activities of the enzymes involved in melatonin synthesis and the contents of two melatonin precursors suggests that swimming depresses pineal melatonin content by enhancing melatonin efflux from the gland.

In young adult male rats bearing a donor anterior pituitary gland grafted for 3 weeks under a kidney capsule, serum prolactin (PRL) concentrations were elevated and exhibited a rhythm with the highest values in the light phase. Serum PRL in control animals did not exhibit a significant rhythm. Eutopic pituitary PRL content, manifesting a biphasic (12-hr) rhythm with crests during the day and night in controls, exhibited a similar pattern in grafted rats though an overall reduction in pituitary PRL content was seen in the grafted animals. Neither the normal biphasic serum testosterone rhythm nor the normal 24-hr rhythm (nocturnal surge) of pineal N-acetyltransferase activity and melatonin content were altered in the hyperprolactinemic rats. Serum thyroxine (T4) and triiodothyronine (T3) and their free indices (FT4I, FT3I) and serum thyrotropin (TSH) were highest during the day in controls and grafted rats and a 12-hr rhythmic component was detected in data for these variables. In the grafted animals, the 12-hr component was reflected in an additional peak at night detectable by testing of means. The overall serum T4, FT4I, and TSH levels were lower in grafted rats though overall T3 and FT3I levels did not differ between grafted and controls. T3 uptake (T3U) values were similar between controls and grafted rats, in both cases exhibiting a fall during the night. Changes in serum thyronines could not be explained by changes in serum binding as assessed by the T3U, and thus may represent changes in thyroidal secretion of T4.(ABSTRACT TRUNCATED AT 250 WORDS)

The effects of crude extracts of bovine, rat, and human pineal glands on prolactin (PRL) release were studied using an in vitro system. In addition, the effects of a known pineal constituent, arginine vasotocin (AVT), and crude bovine pineal extract (bPE) on PRL secretion were studied in vivo. Normal male rat hemipituitaries (HP), incubated with bPE (13 mg tissue/HP)released 200%, 150%, and 285% more PRL into the medium than did their corresponding untreated control halves incubated in either Medium 199 alone, hypothalamic extract, or cerebral cortical extract, respectively. HP incubated with either rat (6 mg of tissue/HP) or human (25 mg of tissue/HP) pineal extract released 110% and 75% more PRL, respectively, than did their corresponding untreated control halves. HP exposed to 10 mg tissue eq of either bovine pineal fraction A1 or bovine pineal fraction A3 released 88% and 63%, respectively, less PRL than did their corresponding untreated control halves incubated in Krebs-Ringer Bicarbonate (KRB) medium. Quantitites of melatonin, thyrotropin-releasing hormone (TRH), or estrogen, comparable to those found in the pineal, had no significant effect on PRL secretion in vitro. The iv injection of either bPE (90 mg tissue/rat) or AVT (10 mug/rat) into estrogen and progesterone-treated male rats resulted in a 40% and 138% increase, respectively, in plasma PRL titers, 10 min after injection, over pre-injection control levels. The per cent of increase in plasma PRL levels in these animals was significantly greater than that observed in control rats receiving either saline or cortical extract. The results suggest that crude extracts of pineal glands of three different species contain prolactin-releasing factor (PRF) activity which is probably not due to any endogenous melatonin, TRH, or estrogen that may be present. Conversely, two bovine pineal fractions, A1 and A3, appeared to exhibit prolactin-inhibiting factor (PIF) activity. We have concluded that the pineal gland may serve as an alternate or supplemental source of PRF and/or PIF.

Previously isolated, adult male mice of a wild-derived strain were sham-operated or pinealectomized (pinx) and 14 days later were paired for 15 min, then grouped three per cage for 7 days. During 15 min pairings, mean latency to initiation of fighting was increased twofold and duration of fighting was reduced 35 to 41 percent in pairs comprised of one or both pinx animals. Although pinealectomy failed to influence which pair member initiated fighting, sham males were ranked as dominant in 75 percent of sham-pinx pairs. Pinealectomy reduced adrenal gland weights among isolated animals and increased in general the effects of social subordination in grouped males. Compared to sham subordinates, pinx subordinates experienced adrenal enlargment and twofold greater wounding and had lighter thymus glands and final body weights. Additionally, however, pinealectomy prevented atrophy of seminal vesicles among subordinates. Thus, present results suggest a role of the pineal gland in influencing intermale aggression and linking social stimuli with testis dysfunction in adult house mice.

Adult male and female hamsters (Mesocricetus auratus) were divided into 3 treatment groups: intact, blinded, and intact hamsters which received daily sc injections of 25 microgram melatonin (MEL) at 1700 h. After 10 weeks of treatment, thyroxine (T4), triiodothyronine (T3) and cholesterol levels in both male and female blinded or MEL-treated animals were depressed compared to plasma titers of their respective intact control groups. Plasma thyrotropin (TSH) levels were depressed by MEL treatment but unaffected by blinding in male hamsters; in contrast, blinded or MEL-treated female hamsters had significantly higher TSH levels than the control group. Plasma triglyceride levels were depressed by both blinding and MEL-treatment in males but were unaffected in females.

Adult male Syrian hamsters either placed in a short photoperiod alone or kept in a long photoperiod and given daily afternoon injections of the pineal indole melatonin (25 micrograms) exhibited splenic hypertrophy and extramedullary hematopoiesis in addition to a marked regression in testicular weight. The testicular regression as well as the changes in spleen weight and histology could be prevented if the animals in short photoperiod were either pinealectomized or implanted subcutaneously with a pellet containing 1 mg melatonin. Female Syrian hamsters given afternoon injections of melatonin for 7 or 12 weeks had ovaries devoid of corpora lutea; additionally, these animals had reduced relative spleen weights compared to the control animals. In conclusion, it is apparent that spleen weight varies with the functional status of the gonads. Splenic hypertrophy accompanied by pineal-induced testicular regression in males may be related to splenic extramedullary hematopoiesis.

The daily s.c injection of 25 microgram melatonin (MEL) in oil into adult male hamsters at 7 p.m. (lights on 6 a.m. to 8 p.m.) for 50 days caused involution of the tests, coagulation of gland and seminal vesicles and depression in pituitary prolactin (Prl) levels. Similar injections of MEL given at 9 a.m. completely failed to cause regression of the sex organs or a depression in pituitary Prl levels. Injections of MEL in the p.m. were completely ineffective in inhibiting either the growth of the gonads and adnexa or the pituitary Prl levels if the animals had been pinealectomized. Likewise, superior cervical ganglionectomy, decentralization of the superior cervical ganglia and anterior hypothalamic deafferetation, procedures which interfere with the sympathetic nerve supply to the pineal gland, negated the ability of p.m. MEL injections to inhibit reproduction in male hamsters. The results indicate that daily MEL injections are capable of suppressing reproductive physiology in male hamsters, but only when the indole is injected late in the light period, in this case, 13 h after light on. The findings also illustrate that daily p.m. MEL injections can inhibit reproduction only in animals that have an intact and sympathetically innervated pineal gland.

Pineal glands and the neurosensory portion of the retinae of adult male rats adapted to a 24-h cycle with lights on from 06.00 to 20.00 h were collected at 9 timepoints during the cycle. Significant rhythms in both pineal and retinal hexosaminidase, beta-glucuronidase, acid phosphatase and beta-glucosidase were observed. For each enzyme, pineals had greater overall activity per unit amount of protein than did retinae. All 4 significantly rhythmic pineal enzymes peaked within 30 min of each other (18.30-19.00 h) whereas the retinal enzymes peaked some 6 h earlier, between 11.30 and 13.45 h. To our knowledge, this is the first report demonstrating 24-h rhythms in lysosomal enzymes in the pineal gland and retina. Since the acrophases (peak times) of these enzymes within each tissue are tightly synchronized yet different between tissues, lysosomes may play unique roles in the physiology of different structures in the photoneuroendocrine system with respect to time in the light-dark cycle.

Blinding adult male golden hamsters was followed by atrophy, within 12 weeks, of the testes and accessory sex organs (seminal vesicles and coagulating glands) and by a significant reduction in pituitary prolactin levels. In experiment 1 blind hamsters received subcutaneously implanted melatonin-beeswax (1:24 mg) pellets at the following intervals: once per week, per 2, 3, 4, 6 weeks, or only one pellet during the 12-week experimental period. The melatonin-beeswax pellets, regardless of the frequency of implantation, overcame completely the inhibitory effects of blinding on reproduction and nearly completely the depressant action of light deprivation on pituitary prolactin levels. In the second study the melatonin-beeswax pellets were implanted subcutaneously into blind hamsters every 2 weeks. The pellets contained either 1 mg, 500, 100, 50, or 1 mug melatonin. With the exception of the 1-mug dosage, melatonin again negated almost totally the inhibitory action of darkness on the gonads and accessory organs and also, for the most part, prevented the drop in pituitary prolactin levels. Based on these studies, when melatonin is chronically administered subcutaneously in a beeswax pellet the minimal dosage of melatonin required to counteract the inhibitory effect of darkness on reproduction seems to be less than 3.6 mug/day. The effects of chronic melatonin treatment are similar to those of pinealectomy.

Blinding resulted in gonadal and prostatic atrophy and reduced plasma thyroxine (T4), free T4 index (FT4I) and reverse triiodothyronine (rT3) levels in adult male hamsters housed in light-to-dark, 14:10 h. Similar effects were seen after daily evening injections of 25 microgram melatonin. Pinealectomy prevented the effects of blinding or melatonin injections. There were no pineal- or melatonin-induced decrements in T3 or thyrotrophin (TSH) concentrations. TSH was elevated by blinding in one experiment but not in another, despite suppression of T4 and FT4I in both. Orally administered melatonin (approximately 245 microgram daily in drinking water through the evening and night) reduced the weight of testes and prostates and slightly lowered plasma T4 and FT4I, indicating the effectiveness of melatonin by this route. The capability of the pineal and of melatonin to suppress plasma T4 is not a result of sex-steroid-induced alteration of plasma binding but is most likely a result of variable suppression of the pituitary-thyroid axis at the level of TSH regulation and also at the level of T4 secretion and/or metabolism. Reduced rT3, but not T3 levels after blinding, may reflect the pineal-induced deficit in T4 as a substrate for rT3 formation, altered peripheral conversion of T4 or altered disposal of thyroid hormones.

The activity of N-acetyltransferase (NAT) and the content of melatonin (MEL) in the rat pineal have been shown to be sensitive to several types of stressors. This study was designed to assess the role of the adrenals in mediating the effect of one such stressor, insulin-induced hypoglycemia, on pineal synthetic activity. Intact and bilaterally adrenalectomized (ADX) adult male rats were kept under light:dark cycles of 14:10 (lights on 0600 h) and injected intraperitoneally with 10 IU insulin at 1300 h, and groups (n = 8) were killed 2, 3, or 4 h postinjection. Plasma catecholamines were assayed by means of high performance liquid chromatography and radioimmunoassay was used to assess pineal NAT activity and MEL content. All injected groups were rendered hypoglycemic by insulin administration. Compared to uninjected controls, plasma epinephrine in hypoglycemic intact rats rose after 2 h, whereas epinephrine did not change in hypoglycemic ADX animals. The increase in epinephrine in intact animals was correlated with a rise in NAT activity at 2 h. Moreover, pineal MEL content at 2, 3, and 4 h was significantly greater than control values. In contrast, no changes in pineal biosynthetic function were found in ADX rats. This differential response by intact and ADX rats suggests that an adrenal product (possibly epinephrine) is responsible for mediating the stimulatory effects of acute insulin-induced hypoglycemic stress on the rat pineal.