Patricia O. Castrillón

Melatonin increases oestradiol-induced bone formation in ovariectomized rats

Abstract: To assess the effect of melatonin on bone metabolism in ovariectomized rats, receiving oestradiol therapy or not, melatonin was administered in the drinking water (25 μg/mL water) and oestradiol (10 μg/kg body weight) or vehicle was given subcutaneously 5 days/week for up to 60 days after surgery. Urinary deoxypyridinoline (a marker of bone resorption) and circulating levels of bone alkaline phosphatase activity (a marker of bone formation), as well as serum calcium and phosphorus levels, were measured every 15 days. Bone area (BA), bone mineral content (BMC), bone mineral density (BMD) and total body fat (expressed as 100 g body weight) were measured by dual‐energy X‐ray absorptiometry at the end of the experiment. Body weight and total body fat were augmented after ovariectomy, and decreased after melatonin or oestradiol treatment. The effect of melatonin on body weight was seen in sham‐operated rats only. Ovariectomy augmented, and melatonin or oestradiol lowered, urinary deoxypyridinoline excretion. This effect of melatonin and oestradiol was seen mainly in ovariectomized rats. The efficacy of oestradiol to counteract ovariectomy‐induced bone resorption was increased by melatonin. Melatonin or oestradiol lowered serum bone alkaline phosphatase activity. Melatonin inhibition was seen mainly on the increase of bone alkaline phosphatase activity that followed ovariectomy. Serum phosphorus levels decreased after melatonin administration and were augmented after oestradiol injection; overall, melatonin impaired the increase of serum phosphorus caused by oestradiol. Ovariectomy decreased, and oestradiol increased, serum calcium levels while melatonin augmented serum calcium in sham‐operated rats only. On day 60 after surgery, BMD and content decreased after ovariectomy and were increased after oestradiol injection. Melatonin augmented BA of spine and BMC of whole of the skeleton and tibia. The highest values observed were those of rats treated concurrently with oestradiol and melatonin. The present results indicate that: (i) melatonin treatment restrained bone remodelling after ovariectomy; (ii) the effect of melatonin required adequate concentrations of oestradiol; (iii) melatonin augmented oestradiol effects on bone in ovariectomized rats; (iv) a counter‐regulation by melatonin of the increase in body fat caused by ovariectomy was uncovered. The melatonin doses employed were pharmacological in terms of circulating melatonin levels but not necessarily for some other fluids or tissues.

Melatonin as a Time-Meaningful Signal in Circadian Organization of Immune Response

Melatonin is synthesized and secreted during the dark period of the light/dark cycle. The rhythmic nocturnal melatonin secretion is directly generated by the circadian clock, located within the suprachiasmatic nuclei in mammals and is entrained to a 24-hour period by the light-dark cycle. The periodic secretion of melatonin may be used as a circadian mediator to any system that can ‘read’ the message. Melatonin seems to act as an arm of the circadian clock, giving a time-related signal to a number of body functions; one of these, the circadian organization of the defense of the organism, is discussed in some detail as an example.

Cerebroventricular Administration of Interferon-Gamma Modifies Locomotor Activity in the Golden Hamster

The present study was undertaken to examine whether the intracerebroventricular (i.c.v.) administration of interferon (IFN)-gamma could modify 24-hour wheel running locomotor activity in the golden hamster. Hamsters implanted with a cannula in the third ventricle received a single i.c.v. injection of 1 µl of murine recombinant IFN-gamma (40 IU/µl) or its vehicle (saline) at ZT 6 or ZT 18 (with ZT 12 defined arbitrarily as the time of lights off) and their activities were monitored during 24 h. The i.c.v. administration of IFN-gamma at ZT 6 produced a significant phase advance in acrophase of rhythm, an effect not seen at ZT 18. Also, IFN-gamma depressed mesor value significantly, the effect was seen at both times. These results clearly showed that the circadian clock could be modified by IFN-gamma microinjections. One explanation could be the presence of IFN-gamma receptor in the rat suprachiasmatic nucleus, supporting a direct effect on the central oscillator. However, another hypothesis could not be ruled out.

Age-Dependent Effect of Pituitary Transplants on Immune Responses in Rat Spleen: Modulatory Effect of Cyclosporine

Male rats were grafted an anterior pituitary within breast muscles on day 5 or under the kidney capsule on day 30 or 60 of life. On the 70th day of life (rats operated on the 5th or 30th day) or on the 100th day of life (rats operated on the 60th day), rats were injected subcutaneously with Freunds complete adjuvant, being killed 2 days later. Rats that had received a pituitary graft on the 30th day showed a greater degree of hyper-prolactinemia than rats grafted on the 5th or 60th day. Analyzed as main factors in a factorial analysis of variance (ANOVA), pituitary transplants augmented splenic natural killer (NK) activity and lipopolysaccharide (LPS)- and concanavalin A (Con A)-induced cell proliferation, and decreased splenic cell number. As indicated by significant interactions between treatment and age of transplantation in a factorial ANOVA, splenic NK activity augmented in rats grafted on the 30th day of life, while LPS and Con A splenic cell proliferation augmented in rats grafted neonatally. Spleen cellularity decreased after pituitary transplants in 30- and 60-day-old rats. In a second study, the effect of cyclosporine on spleen immune responses was tested by administering cyclosporine (5 mg/kg) or vehicle to rats grafted as in experiment 1 for 5 days before sacrifice. Cyclosporine decreased splenic NK activity and LPS- and Con A-induced cell proliferation regardless of the presence of a pituitary graft. In rats grafted on the 30th day of life, cyclosporine reversed the effect of pituitary grafts on splenic NK activity, and ectopic pituitary augmenting NK activity in vehicle-treated rats while decreasing it in cyclosporine-injected rats. Cyclosporine reversed the inhibitory effect of pituitary transplants on spleen cell number. The high circulating prolactin levels found in rats with pituitary grafts were decreased by cyclosporine administration. The results are compatible with age-dependent promoting and inhibitory effects of hyperprolactinemia on the immune responses of the spleen, which were antagonized by cyclosporine immunosuppression.

Age-dependent effect of pituitary transplants on immune responses in rat submaxillary lymph nodes: modulatory effect of the autonomic nervous system.

An anterior pituitary was grafted into the breast muscles of male rats on day 5, or under the kidney capsule on day 30 or 60 of life. At the 70th day of life (rats operated at the 5th or 30th day) or at the 100th day of life (rats operated at the 60th day), rats were injected subcutaneously with Freunds complete adjuvant, being killed 2 days later. Rats pituitary-grafted at the 30th day showed a greater hyperprolactinemia than rats grafted at the 5th or 60th day. Submaxillary lymph node natural killer (NK) activity decreased in neonatally pituitary-grafted rats and increased in rats grafted at the 30th or 60th day of life, while lymph node cellularity decreased in rats grafted at the 30th or 60th day. In the case of lipopolysaccharide (LPS)-and concanavalin A (Con A)-induced cell proliferation in lymph nodes, the only significant factor detected was age of transplantation, the effect being less evident in older animals. To examine whether the autonomic denervation of submaxillary lymph nodes affected immune responses in pituitary-grafted rats, animals were subjected to a unilateral superior cervical ganglionectomy (Gx) and/or a parasympathetic decentralization (Dc; by chorda tympani section), 10 days before immunization with Freunds adjuvant. A unilateral Gx blunted the stimulation of lymph node NK activity in rats receiving a pituitary transplant at the 30th or 60th day, but not at the 5th day. Only in sympathetically denervated lymph nodes a significant effect of pituitary transplants on LPS mitogenic effect was found, with significantly less effect of LPS in rats pituitary-grafted at the 5th or 30th day of life. Regarding Con A, a unilateral Gx or unilateral Gx plus Dc uncovered a significant depressive effect of pituitary transplants with a significantly smaller Con A mitogenic effect at each studied age in ipsilaterally Gx lymph nodes, and at 30 and 60 days of age in ipsilaterally Gx plus Dc lymph nodes. Pituitary grafts augmented Con A- induced proliferation in submaxillary lymph nodes at the 5th day of life while they decreased it at the 60th day of life. Pituitary transplants augmented cellularity at the sympathetically denervated lymph nodes and decreased it at the contralateral sham-operated side. Pituitary transplants also diminished lymph node cellularity at the Dc side, the effect being significant in rats transplanted at the 30th day of life. The results are compatible with age-dependent, inhibitory as well as promoting activities of hyperprolactinemia on immune responses in submaxillary lymph nodes, depending in part on intact autonomic nerves.

Changes in Substance P Content at the Hypothalamic-Pituitary Axis during the Wallerian Degeneration of Peripheral Sympathetic Neurons after Superior Cervical Ganglionectomy in Male Rats: Effect of Hyperprolactinemia

The effects of Wallerian degeneration of the peripheral sympathetic neurons projecting to the hypothalamus on the mechanism of interaction between prolactin and substance P (SP) were examined. The effects of superior cervical ganglionectomy (SCGx) on SP content in various hypothalamic regions and in the hypophysis were evaluated in control and hyperprolactinemic rats. Male rats that received pituitary transplants at the age of 5 days and age-matched sham-operated controls were used. Pituitary grafting significantly increased circulating values of prolactin, as did SCGx. In hyperprolactinemic rats, SCGx partially decreased plasma prolactin levels. Neonatal hyperprolactinemia decreased SP content in the anterior (AH) and posterior (PH) hypothalamus and in the median eminence (ME), but increased it in the mediobasal hypothalamus (MBH). Acute SCGx significantly increased SP in the MBH, PH, and ME. SCGx in hyperprolactinemic animals further increased SP content in MBH. In the ME and Ah, SCGx in pituitary grafted rats decreased SP content as compared with the controls. In the pituitary gland (PG), SCGx only decreased SP content in hyperprolactinemic, but not in control rats. An interaction between peripheral noradrenergic neurons and prolactin to regulate SP within the hypothalamus was positive in the MBH, AH, ME, and PG, but not in the PH. These data indicate the existence of interactive mechanisms between prolactin and the peripheral sympathetic neurons to regulate SP content at the hypothalamic-pituitary axis. Interrelationships between prolactin and SP were also observed.