M. Puig-Domingo

Sexual Dimorphism in N-Acetyltransferase Activity, Hydroxyindole-O-methyltransferase Activity, and Melatonin Content in the Harderian Gland of Syrian Hamsters: Changes following Gonadectomy

Abstract The activities of N-acetyltransferase (NAT) and hydroxyindole-O-methyltransferase (HIOMT) and the melatonin content of the Harderian glands of intact and gonadectomized male and female Syrian hamsters were studied. NAT activity in intact male Harderian glands was twice that of the female. Prepubertal or adult castrated males exhibited a decrease in NAT activity to a level comparable to that seen in the female. Testosterone implants in the castrated males led to a recovery of the original male NAT levels. Intact male hamsters had very low levels of Harderian HIOMT activity and melatonin content in comparison with the glands of the females. Prepubertal gonadectomy but not castration of adult males raised the levels of HIOMT activity and the melatonin content to those of the females. Bilateral ovariectomy had no effect on melatonin content, NAT activity, or HIOMT activity in the female hamster Harderian gland.

Neither the pituitary gland nor the sympathetic nervous system is responsible for eliciting the large drop in elevated rat pineal melatonin levels due to swimming

Since the pineal gland is an end organ of the sympathetic nervous system, stress might increase the synthesis of its hormone, melatonin. The stress of a 10 min swim, which elicits a marked rise in circulating catecholamines, causes a dramatic depression of high pineal melatonin levels at night within 15 min after swimming onset. N-acetyltransferase (NAT) activity is unaffected by the treatment at 15 or 30 min after swimming onset. Within 90 min after initiation of a 15 min swim, high nighttime pineal melatonin levels are restored while NAT values remain elevated. The swimming-induced reduction in high pineal melatonin levels is not influenced by either hypophysectomy, superior cervical ganglionectomy, prazosin (α1-adrenergic receptor blocker) pretreatment, yohimbine (α2-adrenergic receptor blocker) pretreatment, or reserpine (amine depletor) pretreatment. These results indicate that neither hormones secreted from the pituitary gland nor catecholamines secreted from the sympathetic nerves are involved in eliciting the dramatic reduction in elevated pineal melatonin levels in the rat.

Inhibition of Pineal Type-II 5′-Deiodinase Does Not Affect the Nocturnal Increase of N-Acetyltransferase Activity and Melatonin Content in Either Euthyroid or Thyroidectomized Rats

The presence of type‐II thyroxine 5′‐deiodinase (5′‐D) activity in rat pineal gland has been previously described. In the present paper, 5′‐D activity, N‐acetyltransferase (NAT) activity, and melatonin content were measured in the same rat pineal. Each of these constituents exhibits a nocturnal increase with peak values at 0100 h for melatonin (1.20 ± 0.12 ng/gland) and at 0300 h for both 5′‐D (39.5 ± 11.9 fmol/gland/h) and NAT (8.38 ± 1.04 nmol/gland/h) activities. In vivo treatment with iopanoic acid (IOP) completely prevented the nocturnal increase in 5′‐D activity (14.1 ± 2.6 fml/gland/h at 0300 h) with no modification in either the NAT activity or melatonin content. Thyroidectomy greatly enhanced the 5′‐D activity during the dark period (102.9 ± 10.2 vs. 31.6 ± 4.2 fmol/gland/h), reaching a peak at 0200 h; thyroidectomy, however, did not affect daytime pineal 5′‐D activity (3.11 ± 0.78 vs. 2.5 + 0.92 fmol/gland/h). Treatment of rats with IOP acid completely inhibited the pineal 5′‐D activity in both control (7.86 ± 0.88 fmol/gland/h) and thyroidectomized animals (2.24 ± 1.10 fmol/gland/h) with no change in the melatonin content of the gland (1.21 ± 0.32 vs. 0.99 ± 0.18 ng/gland).

Differential responses of rat pineal thyroxine type II 5′-deiodinase and N-acetyltransferase activities to either light exposure, isoproterenol, phenylephrine, or propranolol

Summary1.Compared to pinealN-acetyl transferase (NAT) activity, which exhibited a dramatic drop following acute light exposure at night, nocturnal rat pineal thyroxine type II 5′-deiodinase (5′-D) activity was minimally influenced by the same light exposure. The injection of cycloheximide, a potent inhibitor of protein synthesis, although it did curtail the rise in NAT activity for at least 2 hr, did not elicit decreases in the activities of either 5′-D or NAT enzymes. Propranolol, aβ-adrenergic blocker, either delayed the continued nocturnal rise in 5′-D activity when injected at 0000 hr or slightly enhanced the fall in 5′-D activity when injected at 0200 hr. These results suggest that interruption of the synthesis of proteins is responsible for the slow deterioration of 5′-D activity induced by either light or propranolol.2.The slight fall in 5′-D activity induced by light at night was prevented by isoproterenol; phenylephrine, however, did not prevent the fall and the effect of isoproterenol + phenylephrine was similar to that obtained with isoproterenol alone. On the other hand, the light-inhibited NAT activity recovered after the injection of isoproterenol; phenylephrine did not elicit any effect, but the injection of both isoproterenol and phenylephrine simultaneously caused a greater NAT response than that induced by isoproterenol alone.3.When injected during the day, phenylephrine had no effect on either pineal 5′-D or NAT activities; however, the injection of either isoproterenol alone or isoproterenol + phenylephrine elicited 5-fold and 10-fold increases in nocturnal, light-suppressed 5′-D and NAT activities, respectively. During the day, phenylephrine did not potentiate the effects of isoproterenol on NAT activity as it did at night. When the effects of isoproterenol on the 5′-D activity were compared to rats exposed to light during the day and at night, the activity of 5′-D reached a higher level at night than during the day.

Harderian Gland N-Acetyltransferase Activity in the Male Syrian Hamster: Effects of Gonadectomy, Short Photoperiod Exposure, or Subcutaneous Melatonin Implants

The activities of NAT and HIOMT and the melatonin concentration in the Harderian glands of intact, gonadectomized, and gonadally-regressed male Syrian hamsters were studied. To produce gonadal regression, hamsters were exposed to either artificial or naturally short photoperiods. NAT activity of castrated and gonadally-regressed hamsters was always less in comparison to that of animals with intact gonads. Castrated hamsters exposed to long days showed higher NAT activity than that of castrated animals exposed to short photoperiods indicating that light may have some influence on Harderian NAT independent of the gonadal status. Also, gonadally-regressed hamsters exposed to long photoperiods exhibited higher NAT activity in comparison to gonadally-regressed animals exposed to short days. The HIOMT activity and melatonin content of Harderian glands in all these groups of male Syrian hamster were very low.

Nocturnal increase in the sensitivity of the Syrian hamster pineal gland to isoproterenol is darkness dependent.

Abstract The sensitivity of the Syrian hamster pineal gland to stimulation by isoproterenol is greatly increased in the latter half of the daily dark phase. This increased sensitivity requires a period of dark exposure for up to 6.5 hr. Also, if dark-maintained hamsters are exposed to light in the latter half of the night pineal melatonin levels drop precipitously but can be restimulated by isoproterenol administration. As the interval of light exposure continues, however, the pineal Sensitivity to isoproterenol decreases.

Rhythms in Pineal Immunoreactive Somatostatin in the Syrian Hamster, Mouse, and Gerbil

Immunoreactive somatostatin (IRS) has been previously demonstrated in the pineal gland of different rodent species, and we observed a 24‐hr rhythm in rats. Recent data suggest that the peptide may represent a neurotransmitter in the so‐called peptidergic nerves of the central, pinealopetal innervation of the epiphysis, which may modulate the activity and secretion of the gland. We investigated whether 24‐hr changes of pineal IRS content occurred in Syrian hamsters, gerbils, and mice. Adult males, kept in a 14:10 LD photoperiod, were decapitated at 4‐hr intervals throughout a 24‐hr period. Pineals and median eminences were analyzed for IRS by radioimmunoassay. No significant changes in the median eminence content of IRS with time was observed. As previously described in rats, a statistically significant rhythm of IRS was observed in the pineal of hamsters and mice, with a peak at 2000 hr (mice 51.7 ± 5 pg/pineal; hamsters 26.3 ± 4.6) and a nadir at 2400 hr (mice 30.8 ± 1.4) or 0400 hr (hamsters 8.6 ± 1). However, in the gerbil pineal IRS content remained unchanged throughout the period of study. Since the three species examined have very different melatonin cycles, it is suggested that the melatonin and IRS rhythms are unrelated and independently regulated events within the pineal gland.

Melatonin and 6-methoxy-2-benzoxazolinone (6-MBOA) alter the response of the male Syrian hamster to natural photoperiod

Adult male hamsters bearing either a blank beeswax, 6-methoxy-2-benzoxazolinone (6-MBOA), or melatonin pellet were exposed to 8 weeks (Oct. 6–Dec. 6) of natural autumn decreasing photoperiod (<11 h light) and temperature conditions (mean 10°C for last 4 weeks) or to a 14 h light/10 h dark (14L∶10D) photoperiod and controlled temperature (20°C). Melatonin but not 6-MBOA pellets partially prevented the combined effects of short photoperiod and cold temperatures on the testes and accessory organs. However, both 6-MBOA-and melatonin-treated hamsters maintained outdoors had significantly higher pituitary follicle stimulating hormone (FSH) values compared to their respective indoor-treated controls or to the animals kept outdoors and treated with a blank beeswax pellet. When one compares the various effects of 6-MBOA and melatonin (2 mg/month) on the reproductive system of the male hamster, 6-MBOA is not as effective as melatonin in altering reproductive responses to short photoperiod and cool temperatures at the dose administered.

Identification of immunoreactive somatostatin in the rat Harderian gland: regulation of its content by growth hormone, beta-adrenergic agonists and calcium channel blockers

Immunoreactive somatostatin (IRS) was identified in the male rat Harderian gland (HG) by radioimmunoassay. Tissue was extracted and a displacement curve performed; there were no significant differences between values obtained with serial dilutions of extracted tissue and those from purified somatostatin standard used in the radioimmunoassay. Basal values of HG-IRS were found to be in the nanomolar range (10.8 +/- 3.5 ng IRS/mg protein). Hypophysectomy did not change the HG-IRS but, in vivo growth hormone (GH) treatment led to a dramatic increase (6-7-fold) in the levels of IRS in the HG. Isoproterenol, a beta-adrenergic agonist, when administered in vivo significantly decreased the HG-IRS content. The effect of two different calcium channel blockers on the isoproterenol-induced decrease of HG-IRS was studied; no changes were observed with nifedipine but verapamil, injected one hour after isoproterenol administration, prevented the drop in HG-IRS levels. These data demonstrate the existence of IRS in a new location, the rat Harderian gland, and support a classical endocrine regulation for its tissue concentration.