Pilar Fernández-Mateos

Melatonin normalizes clinical and biochemical parameters of mild inflammation in diet-induced metabolic syndrome in rats

The objective of this study was to evaluate the efficacy of melatonin to affect mild inflammation in the metabolic syndrome (MS) induced by a high‐fat diet in rats. Adult Wistar male rats were divided into four groups (n = 16/group): (i) control diet (3% fat); (ii) high‐fat (35%) diet; (iii) high‐fat diet + melatonin; and (iv) melatonin. Rats had free access to high‐fat or control chow and one of the following drinking solutions for 10 wk: (a) tap water; (b) 25 μg/mL of melatonin. Plasma interleukin (IL)‐1β, IL‐4, IL‐6, IL‐10, tumor necrosis factor (TNF)‐α, interferon (IFN)‐γ, and C‐reactive protein (CRP) were measured at two time intervals, that is, the middle of daylight period and the middle of the scotophase. In addition, a number of somatic and metabolic components employed clinically to monitor the MS were measured. Melatonin decreased the augmented circulating levels of IL‐1β, IL‐6, TNF‐α, IFN‐γ, and CRP seen in obese rats and restored the depressed levels of IL‐4 and IL‐10. Rats fed with the high‐fat diet showed significantly higher body weights and augmented systolic blood pressure from the third and fourth week onwards, respectively, melatonin effectively preventing these changes. In high‐fat‐fed rats, circulating low‐density lipoprotein‐cholesterol, total cholesterol, and triglyceride concentration augmented significantly, melatonin being effective to counteract these changes. Melatonin‐treated rats showed a decreased insulin resistance, the highest values of plasma high‐density lipoprotein‐cholesterol, and the lowest values of plasma uric acid. The results indicate that melatonin is able to normalize the altered biochemical pro‐inflammatory profile seen in rats fed with a high‐fat diet.

Piribedil affects dopamine turnover in cochleas stimulated by white noise

The presence of dopamine (DA) within the cochlea has been previously reported, indicating that its turnover increases under noise stimulation. In the present report, piribedil, a dopaminergic D2 agonist, was used in order to provide evidence of the activity of D2 receptors in the turnover of DA under noise stimulation. Long-Evans rats were intraperitoneally injected with distilled water or with a solution of piribedil one hour previously to either noise or silence exposure. Noise stimulation was performed in an anechoic chamber at 70, 90 or 110 dB SPL for one hour. The animals were then sacrificed and the cochlear contents of DA and its metabolites dihydroxyphenylacetic acid (DOPAC) and homovanillic acid (HVA) were quantified by HPLC with electrochemical detection. The administration of piribedil to animals kept in silence did not modify the cochlear DA, DOPAC and HVA content. Noise stimulation resulted in a decrease of the cochlear DA content and an increase of the cochlear DOPAC and HVA contents in vehicle treated animals. The administration of piribedil resulted in a blockade of this noise induced cochlear DA turnover. These results suggest that piribedil stimulates cochlear D2 receptors controlling the cochlear DA release. Piribedil action on D2 receptors could explain the improvement observed in some cochleo-vestibular diseases signs after piribedil treatment.

Cadmium as an endocrine disruptor: Correlation with anterior pituitary redox and circadian clock mechanisms and prevention by melatonin

To examine the effect of a low dose of cadmium (Cd) as an endocrine disruptor, male Wistar rats received CdCl2 (5ppm Cd) in drinking water or drinking water alone. After 1 month, the rats were euthanized at one of six time intervals around the clock and the 24-h pattern of adenohypophysial prolactin (PRL) synthesis and release, lipid peroxidation, and redox enzyme and metallothionein (MT) gene expression was examined. Cd suppressed 24-h rhythmicity in expression of the PRL gene and in circulating PRL by increasing them at early photophase only, in correlation with an augmented pituitary lipid peroxidation and redox enzyme expression. CdCl2 treatment effectively disrupted the 24-h variation in expression of every pituitary parameter tested except for MT-3. In a second experiment the effect of melatonin (3μg/ml in drinking water) was assessed at early photophase, the time of maximal endocrine-disrupting effect of Cd. Melatonin treatment blunted the effect of Cd on PRL synthesis and release, decreased Cd-induced lipid peroxidation, and counteracted the effect of Cd on expression of most redox enzymes. A third experiment was performed to examine whether melatonin could counteract Cd-induced changes in the 24-h pattern of pituitary circadian clock gene expression and plasma PRL, luteinizing hormone (LH), thyrotropin (TSH), and corticosterone levels. Rats receiving CdCl2 exhibited a suppressed daily rhythm of Clock expression and a significant disruption in daily rhythms of pituitary Bmal1, Per1, Per2, Cry1, and Cry2. The coadministration of melatonin restored rhythmicity in Clock and Bmal1 expression but shifted the maxima in pituitary Per1, Cry1, and Cry2 expression to the scotophase. Melatonin also counteracted the effect of Cd on 24-h rhythmicity of circulating PRL, LH, TSH, and corticosterone. The results highlight the occurrence of a significant endocrine disruptor effect of a low dose of Cd. Generally melatonin counteracted the effects of Cd and ameliorated partially the circadian disruption caused by the pollutant.

Neuroendocrine-immune correlates of circadian physiology: studies in experimental models of arthritis, ethanol feeding, aging, social isolation, and calorie restriction

Virtually all neuroendocrine and immunological variables investigated in animals and humans display biological periodicity. Circadian rhythmicity is revealed for every hormone in circulation as well as for circulating immune cells, lymphocyte metabolism and transformability, cytokines, receptors, and adhesion molecules. Clock genes, notably the three Period (Per1/Per2/Per3) genes and two Cryptochrome (Cry1/Cry2) genes, are present in immune and endocrine cells and are expressed in a circadian manner in human cells. This review discusses the circadian disruption of hormone release and immune-related mechanisms in several animal models in which circulating cytokines are modified including rat adjuvant arthritis, social isolation in rats and rabbits and alcoholism, the aging process and calorie restriction in rats. In every case the experimental manipulation used perturbed the temporal organization by affecting the shape and amplitude of a rhythm or by modifying the intrinsic oscillatory mechanism itself.

Melatonin supplementation decreases prolactin synthesis and release in rat adenohypophysis: correlation with anterior pituitary redox state and circadian clock mechanisms.

In the laboratory rat, a number of physiological parameters display seasonal changes even under constant conditions of temperature, lighting, and food availability. Since there is evidence that prolactin (PRL) is, among the endocrine signals, a major mediator of seasonal adaptations, the authors aimed to examine whether melatonin administration in drinking water resembling in length the exposure to a winter photoperiod could affect accordingly the 24-h pattern of PRL synthesis and release and some of their anterior pituitary redox state and circadian clock modulatory mechanisms. Melatonin (3 µg/mL drinking water) or vehicle was given for 1 mo, and rats were euthanized at six time intervals during a 24-h cycle. High concentrations of melatonin (>2000 pg/mL) were detected in melatonin-treated rats from beginning of scotophase (at 21:00 h) to early photophase (at 09:00 h) as compared with a considerably narrower high-melatonin phase observed in controls. By cosinor analysis, melatonin-treated rats had significantly decreased MESOR (24-h time-series average) values of anterior pituitary PRL gene expression and circulating PRL, with acrophases (peak time) located in the middle of the scotophase, as in the control group. Melatonin treatment disrupted the 24-h pattern of anterior pituitary gene expression of nitric oxide synthase (NOS)-1 and -2, heme oxygenase-1 and -2, glutathione peroxidase, glutathione reductase, Cu/Zn- and Mn-superoxide dismutase, and catalase by shifting their acrophases to early/middle scotophase or amplifying the maxima. Only the inhibitory effect of melatonin on pituitary NOS-2 gene expression correlated temporally with inhibition of PRL production. Gene expression of metallothionein-1 and -3 showed maxima at early/middle photophase after melatonin treatment. The 24-h pattern of anterior pituitary lipid peroxidation did not vary after treatment. In vehicle-treated rats, Clock and Bmal1 expression peaked in the anterior pituitary at middle scotophase, whereas that of Per1 and Per2 and of Cry1 and Cry2 peaked at the middle and late photophase, respectively. Treatment with melatonin raised mean expression of anterior pituitary Per2, Cry1, and Cry2. In the case of Per1, decreased MESOR was observed, although the single significant difference found between the experimental groups when analyzed at individual time intervals was increase at early scotophase in the anterior pituitary of melatonin-treated rats. Melatonin significantly phase-delayed expression of Per1, Per2, and Cry1, also phase-delayed the plasma corticosterone circadian rhythm, and increased the amplitude of plasma corticosterone and thyrotropin rhythms. The results indicate that under prolonged duration of a daily melatonin signal, rat anterior pituitary PRL synthesis and release are depressed, together with significant changes in the redox and circadian mechanisms controlling them. (Author correspondence: danielcardinali@uca.edu.ar; danielcardinali@fibertel.com.ar)

Effect of ethanol on 24-h hormonal changes in prolactin release mechanisms in growing male rats

This study analyzes the effect of chronic ethanol feeding on 24-h variation of hypothalamic-pituitary mechanisms involved in prolactin regulation in growing male Wistar rats. Animals were maintained under a 12:12 h light/dark photoperiod (lights off at 2000 h), and they received a liquid diet for 4 wk, starting on d 35 of life. The ethanol-fed group received a similar diet to controls except that maltose was isocalorically replaced by ethanol. Ethanol replacement provided 36% of the total caloric content of the diet. Rats were killed at six time intervals every 4 h, beginning at 0900 h. Mean concentration of serum prolactin in ethanol-fed rats was 58.7% higher than in controls. Peak circulating prolactin levels occurred at the early phase of the activity span in both groups of rats, whereas a second peak was found late in the resting phase in ethanol-fed rats only. In control rats, median eminence dopamine (DA), serotonin (5-HT), γ-aminobutyric acid (GABA), and taurine levels exhibited two maxima, the major one preceding prolactin release and a second one during the first part of the resting phase. Median eminence DA and 5-HT turnover (as measured by 3,4-dihydroxyphenylacetic acid, DOPAC/DA, and 5-hydroxyindoleacetic acid, 5-HIAA/5-HT ratio) showed a single maximum preceding prolactin, at 0100 h. Ethanol treatment did not affect median eminence DA or 5-HT levels but it decreased significantly their turnover rate. The midday peak in DA and 5-HT levels (at 1300 h) was abolished and the night peak (at 0100 h) became spread and blunted in the ethanol-fed rats. This was accompanied with the disappearance of the 0100 h peak in DA and 5-HT turnover and the occurrence of a peak in 5-HT turnover at 1700 h. Ethanol intake suppressed the night peak in median eminence GABA and taurine (at 0100 h) as well as the midday peak of GABA. Ethanol augmented pituitary levels of DOPAC and 5-HIAA. The results indicate that chronic ethanol administration affects the mechanisms that modulate the circadian variation of prolactin release in growing male rats.

Discontinuous versus Continuous Drinking of Ethanol in Peripubertal Rats: Effect on 24-Hour Pattern of Hypophyseal-Gonadal Axis Activity and Anterior Pituitary Oxidative Stress

Aims: Discontinuous (weekend) consumption of alcohol is common in adolescents and young adults. This study therefore assesses, in peripubertal male rats, the effect of discontinuous as compared to chronic feeding of ethanol or control liquid diet. Methods: Animals received an ethanol liquid diet (6.2 % w/v) starting on day 35 of life. Every week for 5 weeks, the discontinuous ethanol group received the ethanol diet for 3 consecutive days and the control liquid diet for 4 days. At the 5th week, 24 h after the last ethanol administration to the discontinuously ethanol-treated animals, rats were killed at 4-hour intervals beginning at 09.00 h. Chronically administered rats received the ethanol diet until immediately before study. Results: Disrupted 24-hour rhythmicity together with a significant nocturnal increase in plasma luteinizing hormone (LH), testosterone and prolactin (PRL) occurred in the discontinuous ethanol group. Plasma ethanol levels were undetectable at 24 h after the last ethanol treatment. In contrast, after chronic ethanol administration, plasma PRL was increased late in scotophase while LH and testosterone decreased; blood ethanol levels were 2-fold greater than those in discontinuously ethanol-administered rats killed immediately after ethanol withdrawal. Circulating testosterone positively correlated with LH levels in control rats only. Chronic administration of ethanol significantly augmented mean expression of pituitary nitric oxide synthase (NOS)-2, heme oxygenase (HO)-1, Per1 and Per2 genes and disrupted their diurnal rhythmicity. Decreased NOS-1 and NOS-2 expression during scotophase, together with suppression of the rhythm in Per1 and Per2 expression, were found in the discontinuous ethanol group. Conclusions: Abstinence after discontinuous drinking of alcohol in rats, as compared to chronic administration of ethanol, is accompanied by increases of plasma LH and testosterone, a greater PRL response and a less pronounced oxidative damage of the anterior pituitary.

Effect of Subtotal Colectomy on Body Weight and Food Intake in an Experimental Model of Obesity in Male Wistar Rats

The present work analyses the possible influence of the colon in weight gain of obese animals. Subtotal colectomy was performed in rats fed with standard chow or high-fat diets. The results, suggested that subtotal colectomy did not exert any effect on the % of weight gain in rats administered a high-fat diet during the whole experiment as compared to controls. Animals submitted to a high-fat diet during the pre-surgery period and to a standard diet during the post-surgery period gained a lesser % of weight gain as compared to control or high-fat fed rats, together with a decrease in this parameter in colectomized animals. These changes did not agree with average food intake in rats fed with standard or high-fat diets. Surprisingly, the increase in body weight of sham-operated or subtotal colectomized rats fed with high- fat diet cannot be explained by a diminished food intake when compared to controls. Moreover, the change in food intake after surgery showed a correlation with body weight in sham-operated animals, although this correlation disappeared in colectomized rats. Mortality only appeared in colectomized rats administered a high-fat diet. However, stool was normalized and presented normal characteristics when animals returned to be fed with their respective diet after surgery. The results of this study support the concept that the maintenance of a high fat diet may originate a decrease of the intake, overall in subtotal colectomized rats, possibly through neuroendocrine mechanisms related to gut hormones or to adaptive mechanism along the gastrointestinal tract.