Evaluation of the Role of Chronic Daily Melatonin Administration and Pinealectomy on Penicillin-Induced Focal Epileptiform Activity and Spectral Analysis of ECoG in Rats: An In Vivo Electrophysiological Study

In the present study, we aimed to investigate the effects of pinealectomy and chronic melatonin administration on focal epileptiform activity induced by penicillin in the rat cortex and to determine the relation between melatonin levels and electrocorticogram (ECoG) power spectrum. For this purpose, male Sprague-Dawley rats were divided into six groups: control, sham operated, ethanol, melatonin, pinealectomy and pinealectomy + melatonin group. Melatonin-treated rats was intraperitoneally injected with a daily single dose of 10 mg/kg melatonin for 14 days, but the last dose was given 30 min after local application of penicillin as a convulsant agent. Focal epileptiform activity was produced by intracortical administration of penicillin (200 units/1 μl). While chronic melatonin application did not affect either the onset latency or the spike frequency of epileptiform activity, pinealectomy significantly reduced latency to onset of initial epileptiform discharges and increased cortical epileptiform activity. However, acute melatonin administration decreased the epileptiform activity. The results also indicated that exogenously applied melatonin did not change the spectral analysis of ECoG, but pinealectomy led to a reduction in the power of the fast bands (gamma) power in ECoG. We conclude that endogenous melatonin signaling seem to have a tonic inhibitory action on neuronal excitability and epileptiform activity, and also a certain concentration of melatonin required for normal cortical excitability.     

Alteration of tissue zinc distribution and biochemical analysis of serum following pinealectomy in the rat

Eight weeks following pinealectomy in adult male Wistar rats, zinc levels of various tissues were found to be significantly altered: zinc in thoracic aorta was significantly increased, and in serum, pituitary, adrenal, heart, lung, and body hair, it was decreased. Serum biochemical analysis indicated that there was a significant elevation of cholesterol, alkaline phosphatase, sodium, urea, and creatinine in serum from pinealectomised rats. Liver, spleen, and thymus weights were lower following pinealectomy, although hearts were increased. The effects of pinealectomy on zinc levels in serum and tissues and on serum cholesterol and alkaline phosphatase may be related to its effects on vascular reactivity and liver fibrosis.     

Annual testicular cycle of the lizard Anolis carolinensis: effects of pinealectomy and melatonin

The effects of pinealectomy and exogenous melatonin treatment on the reproductive system of male anoles were examined at several different times of year. In September pinealectomy of anoles exposed to either a stimulatory LD 14:10 light cycle or a nonstimulatory LD 10:14 light cycle induced significant testicular growth and development over that observed in sham-operated anoles. At a nonphotosensitive time of year (December) pinealectomy also had a significant progonadal effect but no effect of pinealectomy was seen in February-March. Daily melatonin injections given either in the morning or afternoon (or both) failed to block gonadal growth either (1) in sham-operated or pinealectomized anoles exposed to LD 14:10 in the fall or (2) in pinealectomized lizards exposed to LD 10:14 in the fall. Continuous melatonin administration via subcutaneous silastic implants blocked the progonadal effects of pinealectomy in the winter (December). The results show that pinealectomy can have significant progonadal effects; these effects are seasonal but can encompass phases of the annual testicular cycle which are either photoperiod-dependent or temperature-dependent; and melatonin may be a reproductively active factor involved.     

Pinealectomy increases oxidant damage in kidney and testis caused by hyperthyroidism in rats

Thyroid hormones regulate energy metabolism and act on mitochondria which are an important source of free radicals in the cell. The pineal gland activates antioxidant systems via melatonin secretion and thus has a protective function in body tissues. The present study was conducted to determine the oxidative damage caused by hyperthyroidism in kidney and testis tissues of pinealectomized rats. Experimental animals were allocated to three groups: 1, control group; 2, sham pinealectomy-hyperthyroidic group; and 3, pinealectomy-hyperthyroidic group. Hyperthyroidism was induced by A 3-week intraperitoneal administration of thyroxin after sham pinealectomy or pinealectomy. Malondialdehyde (MDA) and glutathione (GSH) levels were determined in kidney and testis tissues. MDA levels of the kidney and testis tissue in the pinealectomy and hyperthyroidic groups were significantly higher than those in the sham pinealectomy-hyperthyroidic group and the control group (p < 0.001). GSH levels of both kidney and testis tissues were significantly higher in the sham-pinealectomy-hyperthyroidic group when compared to the other two groups (p < 0.001). This increase in GSH levels was more evident in the pinealectomy-hyperthyroidic group than in the control group (p < 0.001). The results of our study demonstrate that MDA and GSH levels in kidney and testis tissues increased due to hyperthyroidism and that pinealectomy made the increase in MDA levels more apparent, while decreasing GSH levels.     

松果体及其褪黑素对大鼠胸腺细胞凋亡的影响

目的探讨松果体及其褪黑素对胸腺细胞凋亡的影响以及Caspase-3的表达。方法选用清洁级SD大鼠,分为正常对照组、假手术对照组、松果体摘除组、松果体摘除 褪黑素腹腔注射7.5mg/kg/d组和松果体摘除 褪黑素腹腔注射15mg/kg/d组。术后4、8周取材。运用TUNEL法检测胸腺细胞的凋亡程度,用ABC法染胸腺Caspase-3阳性细胞,计算机图像分析仪测量阳性细胞面积及其染色强度。以RT-PCR法检测褪黑素干预原代培养胸腺细胞Caspase-3的表达。结果松果体摘除后8周时胸腺细胞凋亡显著增加,补充褪黑素则能明显减少胸腺细胞的凋亡。Caspase-3阳性细胞主要见于胸腺皮质,松果体摘除后胸腺皮质Caspase-3阳性细胞面积增加明显,补充褪黑素则使其下降。褪黑素能上调培养胸腺细胞Caspase-3的表达水平。结论松果体能调控大鼠胸腺细胞的凋亡,松果体摘除促进胸腺细胞的凋亡,补充褪黑素能缓解相关影响。     

Functional relations of the adrenal cortex, thyroid and pineal body. II. Adrenal cortex reaction following epiphysectomy and administration of melatonin

Histologic-cytological and morphometrical changes were investigated in the adrenal cortex of male Wistar-rats following pinealectomy and application of melatonin in eu-, hypo-, and hyperthyroid situations. A rat experiment (at an average of 45 d) to find a possible functional connection between the pineal gland and the adrenal cortex was carried out. In the literature, there are only a few of informations about the role of the pineal in regulating ACTH secretion. The results are very contrarily. We found that pinealectomy is connected with a progressive transformation and melatonin with a little regressive transformation in the adrenal cortex. But, it is not evident, that the glomerular zone is activated after both pinealectomy and application of melatonin. In our opinion, the glomerular zone and the secretion of aldosterone increased after as well pinealectomy as melatonin. Application of melatonin diminishes the function of the pineal gland (see group 4-pinealectomy plus melatonin-where was found a progressive transformation). Under these experimental conditions, one can speak of a "pharmacological pinealectomy" after application of melatonin alone. However, the effect of melatonin on the fascicular zone and the glomerular zone is different. The effects of pinealectomy or application of melatonin in combination with methylthiouracil or thyroxin are relatively unimportant.     

Changes in the learning ability of epiphysectomized rats

The experiments on rats have shown changes in the learning ability of animals. The learning of locomotor reactions of escape and avoidance conditioned reflex was accelerated in animals after pinealectomy. Training of rats in U-shaped and 16-door mazes was significantly affected after pinealectomy. The index of reflex retention (1 and 7 days later) was not changed in the operated animals. Epithalamin (0.5 mg/kg for 5 days) selectively facilitated learning of visual and space discrimination.     

Effects of pinealectomy on the ultrastructure of the golden hamster parathyroid gland

Ultrastructural changes of the parathyroid glands of pinealectomized golden hamsters were investigated. The main changes in the parathyroid glands 1 hour and 1 day after pinealectomy compared with the control and sham-operated groups were an increase of the Golgi complexes, cisternae of the granular endoplasmic reticulum and large vacuolar bodies. In addition, many chief cells contained numerous prosecretory granules in the Golgi areas and many secretory granules in the peripheral cytoplasm. The morphology of the parathyroid glands 7 and 30 days after pinealectomy resembled that of the control parathyroid glands. These results suggest that pinealectomy affects the secretory activity of the parathyroid gland.     

CHANGES IN SEROTONIN LEVEL AND TURNOVER IN DISCRETE HYPOTHALAMIC NUCLEI AFTER PINEALECTOMY AND MELATONIN ADMINISTRATION TO RATS

The influence of the pineal gland on the hypothalamic serotonergic function was examined by studying the effects of long-term pinealectomy (1 month) and melatonin replacement (500 μg/kg; 10 days) on serotonin (5-HT) and 5-hydroxyindoleacetic acid (5-HIAA) content as well as on the in vivo 5-HT synthesis rate in discrete hypothalamic nuclei. Pinealectomy was followed by a significant decrease of 5-HT content in the anterior hypothalamic nuclei (AHN) and the ventromedial hypothalamic nuclei (VMHN), and also in 5-HIAA content in lateral (LPON) and medial preoptic nuclei (MPON). The 5-HT synthesis rate, estimated from the accumulation of 5-hydroxytryptophan after blockade of the 1-amino acid decarboxylase activity, were also decreased in the AHN and the paraventricular hypothalamic nuclei (PVHN) of pinealectomized rats. In contrast, an enhanced 5-HT synthesis rate and basal 5-HIAA content were found in the suprachiasmatic nuclei (SCN) after pinealectomy. Daily treatment with melatonin for 10 days reversed most of the effects induced by pinealectomy. Thus, melatonin increased the levels of 5-HT in the AHN and VMHN, and slightly increased the 5-HIAA content in preoptic nuclei. In addition, melatonin increased the 5-HT synthesis rate in the AHN and VMHN, but also in the MPON, VMHN and dorsomedial hypothalamic nuclei (DMHN) where pinealectomy had no effect. By contrast, melatonin treatment did not affect SCN 5-HT synthesis rate, although it decreased 5-HIAA levels. The results demonstrate that melatonin is able to stimulate 5-HT metabolism in most of the hypothalamic areas, but inhibits SCN 5-HT function. Some of the effects of melatonin seems to be exerted by modulating the synthesis of the amine, although melatonin likely also interacts with other regulatory processes of 5-HT function (i.e. release/uptake). The well defined presence of melatonin receptors in the rat SCN, and its absence in other hypothalamic structures, suggest that this may be the mechanism mediating the differential response to endogenous melatonin. Moreover, the larger effect of exogenous melatonin in relation to pinealectomy suggests the presence of melatonin unespecific effects possibly owing to supraphysiological doses. The present findings may be relevant for the mode of action of melatonin and its implication in several endocrine and behavioral functions mediated by serotonergic neurons. Copyright © 1996 Elsevier Science Ltd     

The effect of pinealectomy on plasma vasopressin response to isotonic, hypertonic and hypovolemic treatments in rats supplemented with L-thyroxine

The present study was conducted to determine the effects of basal, isotonic as well as hypertonic and hypovolemic treatments on fluid-electrolyte balance and plasma AVP levels in rats supplemented with L-thyroxine and pinealectomized L-thyroxine. The animals were initially separated into 4 groups: control (n = 24), L-thyroxine treated (n = 24); L-thyroxine + sham-pinealectomy (n = 24) treated and 4-L-thyroxine + pinealectomy (n = 24) treated. L-thyroxine was given for 4 weeks. At the end of the 4-week experimental period, the sub-groups were formed before decapitation, which were classified as unstimulated (n = 6), isotonic (n = 6), hypertonic (n = 6) and hypovolemic (n = 6) stimulation. Plasma AVP, total triiodothyronine (TT3) and total thyroxine (TT4) levels were examined in plasma by RIA. Hematocrit and osmolality levels were also determined. It was found that the TT3 and TT4 levels showed significant increases in L-thyroxine treated groups (P <0.001). Also, plasma AVP levels increased in the group subjected to L-thyroxine treatment. However, this increase was depicted to be significantly more prominent in L-thyroxine + pinealectomy treated group (P <0.001). The results of the present study indicate that L-thyroxine treatment increases the basal and stimulated AVP release, which became more significant in the pinealectomy plus L-thyroxine treatment group. Moreover, the results indicate that AVP response to hypertonic and hypovolemic stimulations does not undergo any change due to supplementation by L-thyroxine treatment and/or pinealectomy plus L-thyroxine.     

Effects of pinealectomy and exogenous melatonin on ghrelin and peptide YY in gastrointestinal system and neuropeptide Y in hypothalamic arcuate nucleus: immunohistochemical studies in male rats

It is reported that the pineal gland and its main hormone melatonin may have a role in the regulation of ghrelin synthesis in the brain. Stomach is the place where ghrelin is predominantly expressed and secreted. One aim of this study was to investigate possible effects of pinealectomy and melatonin treatment on gastric ghrelin amount. The studies on the effects of the pineal gland on leptin and ghrelin arises the question whether the pineal gland has also effects on the other energy-regulatory peptides such as peptide YY (PYY) and neuropeptide Y (NPY). Therefore, we also aimed to investigate the changes in the immunohistochemical staining of intestinal PYY and hypothalamic NPY following pinealectomy and melatonin treatment. Serum PYY levels were also investigated. Sprague-Dawley rats were divided into four groups as sham-operated (SHAM), sham-operated with melatonin treatment (SHAM-MT), pinealectomised (PNX) and melatonin-treated PNX (PNX-MT) groups. The cells immunostained for ghrelin were abundant throughout the gastric mucosa in all the groups. Neither pinealectomy nor exogenous melatonin affected significantly immunohistochemical staining of ghrelin in stomach. Pinealectomy resulted in a significant increase in immunohistochemical staining of PYY in ileum. The results of serum PYY measurement corresponded closely to the data obtained by immunohistochemical analysis of PYY in ileum, being significantly lower and higher in SHAM and PNX groups, respectively. Pinealectomy caused a decrease in NPY synthesis in ARC as understood from low immunohistochemical staining of NPY. Melatonin treatment increased NPY synthesis in SHAM rats and restored reduction in NPY synthesis caused by pinealectomy. In conclusion, the pineal gland and its main hormone melatonin can be suggested to have a role in the regulation of NPY synthesis in ARC and PYY in gastrointestinal system.     

Maternal transfer of photoperiodic information in Siberian hamsters. IV. Peripubertal reproductive development in the absence of maternal photoperiodic signals during gestation

Peripubertal reproductive development of Siberian hamsters is influenced by photoperiodic information received during gestation; the maternal pineal is important for this process. We observed that in the absence of the maternal pineal, the fetus appears to receive no information about gestational photoperiods. This is not the equivalent of receipt of a long-day signal by the fetus. Pinealectomized and sham-operated pregnant females were exposed to photoperiods of 12L:12D, 14L:10D, 16L:8D, or constant light (LL); young were reared from birth to 28 days of age in LL or 14L:10D. Regardless of the gestational photoperiod, LL-reared male young born to pinealectomized dams had smaller testes than LL-reared young of pineal-intact dams exposed to 16L:8D while pregnant. Thus, pinealectomy did not result in transmission of a long-day signal, nor did young born of pinealectomized dams receive short- or intermediate-day signals. Unlike young of pineal-intact females exposed to 12L:12D or 14L:10D while pregnant, young born of pinealectomized dams had small testes when reared in 14L:10D, irrespective of gestational photoperiod. Uterine weights of female young presented similar patterns of responses. In a second experiment, adult females were entrained to 12L:12D, 14L:10D, or 16L:8D for 3 wk prior to pinealectomy to determine if the effect of maternal pinealectomy would be altered. Entrainment to the new photoperiod prior to surgery did not alter the effects of maternal pinealectomy.     

Melatonin and the synthesis of vasopressin in pinealectomized male rats

The pineal hormone, melatonin, is known to modify, under different experimental conditions, neurohypophysial hormone secretion in the rat. The aim of this study was to investigate the effect of melatonin on the vasopressin biosynthesis rate in the hypothalamus of either pinealectomized or sham-operated rats, using the colchicine method. To estimate whether colchicine affects the function of the neurohypophysis in these animals, the neurohypophysial and plasma vasopressin levels were also measured. The vasopressin synthesis rate was increased after pineal removal, when compared with sham-operated animals, and melatonin strongly inhibited the rise in the hormone synthesis due to pinealectomy. After pineal removal plasma vasopressin concentration was significantly elevated, and melatonin attenuated this effect. On the contrary, the neurohypophysial vasopressin content was significantly decreased after pinealectomy, but it was not further modified by melatonin.Thus, melatonin suppresses the synthesis and secretion of vasopressin in pinealectomized rats. The present results confirm our previous reports as to the inhibitory impact of the pineal on both vasopressin synthesis and release and suggest that melatonin may mediate the effect of the pineal gland on vasopressinergic neuron activity.     

The effect of pinealectomy and jejunal loop diversion on rat small bowel crypt cell kinetics

Previously, it was found that diversion of an isolated loop of jejunum into the colon was associated with a significantly diminished crypt cell proliferation rate in the isolated loop, probably principally because of the diminished amount of nutrient available to the diverted mucosa. It has also been shown previously that removal of the pineal gland is associated with a considerable increase in the jejunal crypt cell mitotic rate. In the present investigation it was found that following pinealectomy, whilst the rise in crypt cell proliferation elsewhere in the rat small intestine was maintained at the expected level, the mitotic rate in the crypts of an isolated jejunal loop attached to the colon was also increased to a similar level, despite the fact that this isolated loop was in contact with a considerably diminished level of luminal nutrients. Thus, the expected hypoproliferative effects of jejunal isolation were overridden by the hyperproliferative effects of pinealectomy and the effects of pinealectomy appeared to be independent of the particular changes in luminal environment produced in this experiment. The significance of these findings is discussed in the light of the available literature.     

Seasonal effects of pinealectomy on testicular recrudescence and secretory activity of seminal vesicles in the catfish Heteropneustes fossilis (Bloch)

The effects of pinealectomy on testicular activity and secretory activity of seminal vesicles were examined in the catfish Heteropneustes fossilis under various combinations of photoperiod and temperature during different periods of the annual reproductive cycle. Pinealectomy had no effect on gonadal activity during the preparatory, prespawning and spawning periods of the reproductive cycle. However, during the postspawning period, under long (LD 14:10) or short (LD 9:15) photoperiod at 25° C or at gradually increasing ambient temperature, pinealectomy accelerated testicular recrudescence and secretory activity of the seminal vesicles. Nevertheless. during this period the presence of the pineal facilitated the recrudescence of testes and seminal vesicles in catfish exposed to continuous light (LL), continuous darkness (DD) and 12 hL:l2 hD (LD) at 25° C. These findings suggest that the role of the pineal in catfish reproduction is variable and depends upon the photoperiod and temperature regimes to which the fish are exposed, as well as on the time of the year and the state of the reproductive cycle. The results also suggest that the effects of pinealectomy in catfish are mediated through an influence on the hypothalamo-hypophyseal gonadal axis.     

Influence of the pineal gland on testicular function in offspring of pinealectomized rats

Female Sprague-Dawley rats exposed to a short (6L:18D) photoperiod from 21 days of age were mated when they reached 55 days of age. On Day 2 of gestation animals were pinealectomized or sham-operated. On Day 5 after birth male pups of the two groups of dams were either pinealectomized or sham-operated. They were killed at 42 and 49 days of age. In offspring born to sham-operated dams and in those born to pinealectomized mothers, neonatal pineal ablation resulted in increased testicular testosterone and androstenedione content. In sham-operated and neonatally pinealectomized rats removal of the maternal pineal gland induced a decrease in testicular testosterone and androstenedione content. In contrast, after maternal pinealectomy there was a decrease in plasma testosterone and dihydrotestosterone values and testicular dihydrotestosterone content in sham-operated rats but not in those neonatally pinealectomized. We conclude that (1) the pineal glands of the mother and offspring are required to maintain normal testicular testosterone and androstenedione content in the rat, and (2) the pineal of the offspring influences the inhibitory effects of maternal pinealectomy on testicular dihydrotestosterone content and on plasma testosterone and dihydrotestosterone concentration in the offspring.     

Advanced radiological work-up as an adjunct to decision in early reconstructive surgery in brachial plexus injuries

Background

Although the injury to the peripheral nervous system is a common clinical problem, understanding of the role of melatonin in nerve degeneration and regeneration is incomplete.

Methods

The current study investigated the effects of neonatal pinealectomy on the sciatic nerve microarchitecture in the chicken. The chickens were divided into two equal groups: unpinealectomized controls and pinealectomized chickens. At the end of the study, biochemical examination of 10 sciatic nerve samples from both groups was performed and a quantitative stereological evaluation of 10 animals in each group was performed. The results were compared using Mann-Whitney test.

Results

In this study, the results of axon number and thickness of the myelin sheath of a nerve fiber in newly hatched pinealectomy group were higher than those in control group. Similarly, surgical pinealectomy group had significantly larger axonal cross-sectional area than the control group (p < 0.05). In addition, the average hydroxyproline content of the nerve tissue in neonatal pinealectomy group was higher than those found in control group. Our results suggest that melatonin may play a role on the morphologic features of the peripheral nerve tissue and that melatonin deficiency might be a pathophysiological mechanism in some degenerative diseases of peripheral nerves. The changes demonstrated by quantitative morphometric methods and biochemical analysis has been interpreted as a reflection of the effects of melatonin upon nerve tissue.

Conclusion

In the light of these results from present animal study, changes in sciatic nerve morphometry may be indicative of neuroprotective feature of melatonin, but this suggestion need to be validated in the human setting.     

Diurnal Variations in [125I]Melatonin Binding by Rat Thymus Membranes: Effects of Continuous Light Exposure and Pinealectomy

Binding of melatonin by rat thymus membranes exhibited diurnal changes. Binding increased during the daytime and reached maximal values before entering the dark period. Then, binding decreased rapidly during the dark phase. In rats kept in light at night, binding of [¹²⁵I]melatonin by membranes was significantly higher than in animals that entered the normal dark period. Neonatal pinealectomy, which suppresses the circadian rhythm of melatonin, led to an increase in melatonin binding of 106%. Moreover, in animals maintained under continuous light exposure, which corresponds to functional pinealectomy, binding of melatonin by thymus membranes also increased in a time-dependent manner. The results support the hypothesis of a regulatory role of melatonin in the thymus in which melatonin downregulates its own binding sites.     

Effects of melatonin and epiphysectomy on electrical activity of the rat brain at different times of the day]

The pineal hormone melatonin in a dose-effect manner (0.1, 1 and 10 mg/kg) reorganized EEG spectral characteristics of rat sensorimotor cortex and hippocampus, decreasing medium (6-12 Hz) and high frequency (12-22 Hz) waves only in the evening. On the contrary, pinealectomy increased high-frequency waves only during morning hours. It is suggested that the observed changes may accompany anxiolytic and hypnotic effects of melatonin.