David E. Blask

EFFECTS OF MELATONIN ON THE CELL CYCLE KINETICS AND ESTROGEN-RESCUE OF MCF-7 HUMAN BREAST CANCER CELLS IN CULTURE

Cos S, Blask DE, Lemus‐Wilson A, Hill AB. Effects of melatonin on the cell cycle kinetics and “estrogen‐rescue of MCF‐7 human breast cancer cells in culture. J Pineal Res 1991:10:3642.

Pineal melatonin inhibition of tumor promotion in the N-nitroso-N-methylurea model of mammary carcinogenesis : potential involvement of antiestrogenic mechanisms in vivo

SummaryTheN-methyl-N-nitrosurea (NMU) model of hormone-responsive rat mammary carcinogenesis was used to address the hypothesis that melatonin (Mel), the principle hormone of the pineal gland, inhibits tumorigenesis by acting as an anti-promoting rather than an anti-initiating agent. Daily late-afternoon injections of Mel (500 μg/day), restricted to the initiation phase of NMU mammary tumorigenesis, were ineffective in altering tumor growth over a 20-week period. When Mel treatment was delayed for 4 weeks after NMU and then continued through the remainder of the promotion phase, only tumor number was significantly lower than in controls. However, when Mel injections encompassed the entire promotion phase, both tumor incidence and number were significantly lower than in the controls. Although elimination of the endogenous Mel signal via pinealectomy promoted tumor growth, the effect was not statistically significant. Serum levels of estradiol and tumor estrogen receptor content were unaltered by either Mel or pinealectomy. While Mel treatment failed to affect circulating prolactin levels, pinealectomy caused a two-fold increase in serum prolactin. The estradiol-stimulated recrudescence of tumors following ovariectomy was completely blocked by either 20, 100 or 500 μg Mel/day or tamoxifen (20 μg/day). Thus, Mel appears to be an antipromoting hormone that may antagonize the tumor-promoting actions of estradiol in this model of mammary tumorigenesis.

Inhibitory effects of the pineal hormone melatonin and underfeeding during the promotional phase of 7,12-dimethylbenzanthracene-(DMBA)-induced mammary tumorigenesis

The effects of melatonin (Mel) and/or underfeeding (30% food restriction) on 7,12-dimethylbenzanthracene (DMBA)-induced mammary tumorigenesis were examined in female Sprague-Dawley rats fed a semipurified diet. During the promotional phase of tumorigenesis, the animals began receiving either daily afternoon Mel (250μg) or saline vehicle injections s.c. for 15 weeks. As compared with fed animals, underfed rats had a lower tumor incidence, tumor number and size while the latency to onset and regression of tumors was increased. Melatonin in fed rats moderately suppressed tumor incidence and number. However, the combination of Mel treatment and underfeeding caused the most marked inhibition of tumorigenesis as compared with either treatment alone. These results indicate that Mel administration and/or underfeeding during the promotional phase inhibit DMBA-induced mammary tumorigenesis perhaps via neuroendocrine and/or peripheral endocrine mechanisms.

Effects of the pineal hormone melatonin on the anchorage-independent growth of human breast cancer cells (MCF-7) in a clonogenic culture system.

Only physiological levels of melatonin exert an antiproliferative effect on MCF-7 breast cancer cells grown in an anchorage-dependent culture system. We investigated melatonins effect on the anchorage-independent growth of MCF-7 cells as well as the dose-response characteristics of this indoleamine under clonogenic culture conditions. Melatonins inhibitory effect, with respect to the number and size of colonies formed, exhibit a linear dose-response curve with pharmacological concentrations producing a maximal inhibition while subphysiological levels of melatonin induce minimal inhibition. These results indicate that cellular attachment may modify the sensitivity of MCF-7 cells towards melatonin.

Hypothalamic site of action of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD)

Administration of TCDD produced a significant decrease in the serum concentration of prolactin (PRL) detected in rats after 4 hr compared to pair-fed vehicle controls and noninjected controls. This effect of TCDD was reversed by pimozide, a dopamine receptor antagonist. These data suggest that TCDD decreased the release of PRL from the adenohypophysis either by a direct effect on the gland or by altering the dopamine concentration in the median eminence (ME). Concentrations of TCDD from 5 to 500 ng/ml had no direct effect on the ability of the adenohypophysis to secrete PRL in vitro. However, the dopamine concentration increased to 3.24 +/- 0.07 ng per ME in TCDD-treated rats compared to 2.81 +/- 0.08 ng in vehicle controls. This is a dramatic alteration in the dopamine concentration, since the dopamine is being measured in the portal circulation which exhibits a rapid turnover. The rate constant of dopamine depletion after alpha-methyl-p-tyrosine and the turnover rate were also significantly elevated in the ME of TCDD-treated rats. These data provide the first biochemical evidence for a hypothalamic site of action of TCDD. Since dopamine is inhibitory to PRL release from the adenohypophysis, increased ME steady-state concentrations and turnover of this catecholamine may be responsible for the decreased concentration of serum PRL detected within 4 hr of TCDD injection. Thus, one of the early modes and sites of action of TCDD is to elevate the dopaminergic activity of the tuberoinfundibular nucleus. A hypothalamic site of action for TCDD may result in a number of the endocrinological effects known to be produced by exposure to TCDD.

Antigonadotrophic and Prolactin-Inhibitory Effects of Melatonin in Anosmic Male Rats

Immature (25–26 days of age) male rats housed on 14 h of light per day (lights on 06.00–20.00 h) were either olfactory bulbectomized, rendering them anosmic, or left intact. On the day following bulbe

Prolactin Cell Activity in Female and Male Syrian Hamsters: An Apparent Sexually Dimorphic Response to Light Deprivation and Pinealectomy

In order to determine the role of the pineal gland in mediating the effects of long-term light deprivation on prolactin (PRL) cell activity in a highly photosensitive species, PRL synthesis, storage and release were determined in both female and male Syrian hamsters that were either blind, blind and pinealectomized or left intact for 14 weeks. PRL release was determined in vivo by measuring the amount of immunoreactive (RIA) PRL in the serum of the animals in each group with a heterologous RIA for hamster PRL. Binding resulted in a 98 and 88% reduction in serum PRL levels in female and male hamsters, respectively. Pinealectomy largely prevented the suppressive effects of blinding on PRL release; however, PRL levels in blind pinealectomized animals were intermediate between those in intact and blind animals. PRL synthesis was evaluated by assessing the amount of 3H-leucine incorporated into PRL by anterior pituitaries in vitro. In female hamsters 14 weeks of light deprivation resulted in an 87% decrease in the incorporation of 3H-leucine into newly synthesized PRL whereas in males only a 40% reduction occurred. While pinealectomy completely prevented the inhibitory effects of blinding on PRL synthesis in males, it was less effective in female hamsters inasmuch as PRL synthesis was still nearly 50% lower in blind pinealectomized animals than in controls. Stored PRL, as represented by the total amount of RIA-PRL in vitro, was 96% lower in blind female hamsters as compared with intact controls; in blind male hamsters, stored PRL was reduced by 77%.(ABSTRACT TRUNCATED AT 250 WORDS)

Evidence for an inhibitory influence of the pineal on prolactin in the female rat.

The effect of the pineal gland on prolactin (PRL) synthesis, storage and release was tested in female rats. To do this, we incubated in vitro hemi-anterior pituitaries from 80-day-old female rats that had been rendered, 8 weeks previously, either blind-anosmic, blind-anosmic and pinealectomized or left intact. Reproductive organ weights, pituitary weights and pituitary DNA content were decreased in animals rendered both blind and anosmic. These effects were reversed by removal of the pineal gland. Additionally, the serum levels of PRL were diminished in blind-anosmic rats from that of intact controls. The synthesis of PRL in vitro was dramatically reduced in the pituitaries of blind-anosmic rats as evidenced by a 63% decrease in [3H]leucine incorporation into PRL from that observed in the intact group. Likewise, the total amount (medium + pituitaries) of radioimmunoassayable PRL in vitro was depressed by 26% in the blind-anosmic group as compared with intact controls. Pinealectomy reversed the reductions in de novo synthesized and total immunoreactive PRL in vitro. From these studies we conclude that in the blind-anosmic female rat the pineal chronically inhibits PRL synthesis and storage in the pituitary and, possibly, its release into the blood. These pineal-induced effects could be accounted for by a reduction in the pituitary mammotroph as might be indicated by the decrease in pituitary DNA content observed in dual-sensory deprived rats.

Melatonin Action on Oncogenesis

Great strides have been made within the last decade in our understanding of the processes involved in oncogenesis not only at the organismal level but at the cellular-molecular level as well. It has become clear that hormones and growth factors play a prominent role in the mechanisms governing oncogenesis. The importance of the hormonal and growth factor regulation of oncogenesis is particularly evident with respect to cancer of the breasts1.

Opioid and dopamine involvement in prolactin release induced by arginine vasotocin and vasopressin in the male rat

The potential involvement of the endogenous opioid and dopamine (DA) systems in the mechanism(s) mediating arginine vasotocin (AVT)- and arginine vasopressin (AVP)-induced prolactin (PRL) release was investigated in vivo. The injection of AVT (5 micrograms) into unanesthetized male rats resulted in a 2-fold stimulation of PRL release 15 min later, followed by an inhibition of PRL release 30 min thereafter; both the stimulatory and inhibitory PRL responses to AVT were obviated by naloxone (NAL) (200 micrograms). Similarly, the administration of either AVT or AVP (5 micrograms) to urethane-anesthetized rats led to a 3- and 5-fold increase in plasma PRL levels, respectively, 10 min after injection. The PRL stimulatory response to both peptides was completely blocked by pretreating the animals with apomorphine (APO) (5 mg); however, the injection of APO by itself had no effect on PRL secretion in these animals. Both AVT and AVP were also effective in stimulating PRL release 10 min after injection in estrogen (50 micrograms)-progesterone (25 mg) (EP)-treated rats anesthetized with urethane. APO negated the PRL stimulatory response to these compounds in the EP-treated rat as well. Normal, urethane-treated rats experienced a 7- to 8-fold increase in PRL levels 20 min following the injection of methysergide (MET) (250 micrograms). Both AVT and AVP caused approximately a 2.5-fold greater PRL response in MET-treated animals than in AVT and AVP controls, respectively; however, only in the MET + AVT-treated rats was the PRL stimulatory response greater than in the MET controls. MET probably stimulated PRL through its DA antagonistic properties.(ABSTRACT TRUNCATED AT 250 WORDS)