Reductions in hamster serum thyroxine levels by melatonin are not altered by changes in serum testosterone.

Daily melatonin injections reduce reproductive and thyroid hormones in male Syrian hamsters. The interrelationship between the decline in these hormones is not known. To explore this relationship, male Syrian hamsters were divided into four groups: castrated, implanted with testosterone (5-mm silastic implants), both treatments, or neither treatment. One-half of each group of hamsters (n = 7 or 8) were injected with melatonin (25 mg) daily at 1730 h. The other half of each group received daily vehicle injections. Ten weeks later, the hamsters were anesthetized and decapitated. Testes weights, serum testosterone, and serum thyroxine levels were measured. As expected, testes and serum testosterone levels were uniformly low in all of the melatonin-treated hamsters. All of the melatonin-treated groups also had lower than normal thyroxine values irrespective of gonadal treatment. Interestingly, in the non-melatonin-treated hamsters, serum thyroxine values were decreased in the castrated group and increased in the testosterone-implanted group. These results suggest that castration can reduce serum thyroxine levels in male Syrian hamsters and that replacement of testosterone restores these levels to normal. Notably, the declines in thyroxine levels produced by daily melatonin injections were not restored by testosterone implants in castrated or intact hamsters. Therefore, melatonin-induced reductions in thyroxine are not mediated by concurrent reductions in serum testosterone levels. It appears that melatonin-induced reductions in serum thyroxine levels do not use the same mechanism as castration-induced reductions.

Acute melatonin and para-chloroamphetamine interactions on pineal, brain and serum serotonin levels as well as stress hormone levels.

para-Chloroamphetamine, an amphetamine analog, alters serotonergic neurochemistry. In previous reports, melatonin (MEL), when administered with other amphetamine analogs, altered the decline in serotonin content produced by these analogs. The present studies assessed the effects of various doses of melatonin and p-chloroamphetamine on serotonin levels in numerous brain regions in male rats. Melatonin (10, 25 or 50 mg/kg, s.c.) and p-chloroamphetamine (3 or 5 mg/kg, s.c.) were administered and, 3 h later, brain samples and serum were collected. Serotonin levels in the serum and various regions of the brain were assayed using high-performance liquid chromatography. Melatonin in combination with a high dose of p-chloroamphetamine (5 mg/kg) produced cumulative deficits in serotonin levels in the serum. However, serotonin levels in the pineal, cortex or brain stem in all combined melatonin and p-chloroamphetamine groups were not significantly different from groups that received p-chloroamphetamine alone. Serum adrenocorticotropin (ACTH) and corticosterone levels were significantly elevated in the melatonin and p-chloroamphetamine combined groups, suggesting that animals receiving both treatments were more stressed than control animals or animals receiving melatonin or p-chloroamphetamine alone. These results indicate that melatonin does not alter p-chloroamphetamine-induced deficits in central serotonin levels. The increased serum adrenocorticotropic hormone, corticosterone and serotonin levels observed following melatonin and p-chloroamphetamine treatment suggest that this combination may have adverse peripheral effects.

Effect of alcohol consumption on adult and aged bone: composition, morphology, and hormone levels of a rat animal model.

To determine the effect of life-long alcohol consumption on the adult and aged rat model, 4-week-old, female Sprague-Dawley rats were divided into three diet groups. Alcohol-treated animals were fed a modified Lieber-DeCarli diet ad libitum containing 35% ethanol-derived calories, whereas the pair-fed animals (weight-matched to ethanol rats) received an isocaloric liquid diet in which maltose-dextrin substituted calories supplied by ethanol. Chow animals were fed a standard rat chow ad libitum. Proximal tibiae (primarily cancellous bone) and femora (primarily cortical bone) were removed for analysis after 3, 6, 9, 12, or 18 months on the diets. Serum was collected for analysis of calcium levels, the calcium regulating hormones; parathyroid hormone, 25-hydroxyvitamin D, calcitonin, corticosterone, estradiol, testosterone, and IGF-1. Creatinine, SGOT/AST, and SGPT/ALT levels were measured to determine kidney and liver integrity. Previous studies, with young animals, showed that chronic alcohol consumption during the age of bone development reduced bone density and bone mass in both cortical and cancellous bone. The present study demonstrates that these reductions last throughout life, whereas morphological values, such as length and diameter, attain control levels. Calcium regulating hormones and sex hormones are essentially normal and do not appear to be the primary causative agent for adult alcohol-induced osteopenia, but it appears to be due to a more direct effect of alcohol on bone cells.

Time-dependent effects of melatonin on immune measurements in male Syrian hamsters.

Adult, male Syrian hamsters received daily subcutaneous melatonin (25 microg) injections or vehicle injections at 08:00 or 17:00 hr for 11 weeks. Body weights were measured weekly throughout the experiment and testes weights, spleen weights, and serum was collected at the end of the experiment. The spleens were sectioned and immunocytochemically analyzed for immunoglobulin G and serum levels of interferon-gamma, interleukin-2, and interleukin-4 were determined in heterologous mouse assays. Melatonin injections at 17:00 hr, but not at 08:00 hr, increased body weights, decreased testes weights and serum testosterone levels, and had no effect on immunoglobulin G content in the spleen. Likewise, melatonin injections at 17:00 hr, but not at 08:00 hr, increased serum interferon-gamma levels, had no effect on interleukin-2 levels, and appeared to increase interleukin-4 levels. Since melatonin injections at 08:00 hr were ineffective in altering immune measurements and correlations between reproductive measures and immune measures were high, the most parsimonious explanation for these results is that melatonin injections at 17:00 hr depressed reproductive hormone levels and these depressed levels altered immune measures.

Immune responsiveness of splenocytes after chronic daily melatonin administration in male Syrian hamsters.

The interrelationships between the immune system and the pineal hormone, melatonin, have been explored recently. The present studies investigated the effects of daily melatonin injections on reproductive and spleen function in male Syrian hamsters. Testes weights and serum testosterone levels were depressed after 8-10 weeks of daily melatonin injections. Melatonin-treated hamsters exhibited increased splenic lymphoproliferative responses to a polyclonal T-cell mitogen (concanavalin A (Con-A)), but decreased proliferation following stimulation with a polyclonal B-cell mitogen (lipopolysaccharide). It appears that daily melatonin injections in male hamsters increase the T-cell-mediated immune capacity while reducing the antibody-mediated immune potential. These data suggest that chronic, daily melatonin alters immune system responsiveness in hamsters by shifting the balance of cellular and humoral reactivity.

The effect of detomidine and its antagonism with tolazoline on stress-related hormones, metabolites, physiologic responses, and behavior in awake ponies.

Six ponies were used to investigate the effect of tolazoline antagonism of detomidine on physiological responses, behavior, epinephrine, norepinephrine, cortisol, glucose, and free fatty acids in awake ponies. Each pony had a catheter inserted into a jugular vein 1 hour before beginning the study. Awake ponies were administered detomidine (0.04 mg/kg intravenously [i.v.]) followed 20 minutes later by either tolazoline (4.0 mg/kg i.v.) or saline. Blood samples were drawn from the catheter 5 minutes before detomidine administration (baseline), 5 minutes after detomidine administration, 20 minutes before detomidine administration which was immediately before the administration of tolazoline or saline (time [T] = 0), and at 5, 30, and 60 minutes after injections of tolazoline or saline (T = 5, 30, and 60 minutes, respectively). Compared with heart rate at T = 0, tolazoline antagonism increased heart rate 45% at 5 minutes. There was no difference in heart rate between treatments at 30 minutes. Blood pressure remained stable after tolazoline, while it decreased over time after saline. Compared with concentrations at T = 0, tolazoline antagonism of detomidine in awake ponies resulted in a 55% increase in cortisol at 30 minutes and a 52% increase in glucose at 5 minutes. The change in free fatty acids was different for tolazoline and saline over time. Free fatty acids decreased after detomidine administration. Free fatty acids did not change after saline administration. After tolazoline administration, free fatty acids increased transiently. Tolazoline tended to decrease sedation and analgesia at 15 and 60 minutes postantagonism. Antagonism of detomidine-induced physiological and behavioral effects with tolazoline in awake ponies that were not experiencing pain appears to precipitate a stress response as measured by cortisol, glucose, and free fatty acids. If antagonism of an alpha-agonist is contemplated, the potential effect on hormones and metabolites should be considered.

Alcohol consumption inhibits bone growth and development in young actively growing rats.

Adolescence is an age of widespread alcohol abuse, but the effect of alcohol consumption on bone formation has not been studied in the young population. This study addresses the effect of alcohol on the early phases of bone growth and development in an animal model. Four-week-old, female Sprague-Dawley rats were divided into three groups. Alcohol-treated animals were fed a modified Lieber-DeCarli diet ad libitum containing 35% ethanol-derived calories, whereas the pair-fed animals (weight-matched to ethanol rats) received an isocaloric liquid diet in which maltose-dextrin substituted calories supplied by ethanol. Chow animals were fed a standard rat chow ad libitum. Proximal tibiae (primarily cancellous bone) and femora (primarily cortical bone) were removed for analysis after 2, 4, 6, or 8 weeks on the diets. Serum was collected for analysis of calcium levels, osteocalcin, corticosterone, growth hormone, parathyroid hormone, and 25-hydroxyvitamin D. The most rapid weight gain occurred between 6 and 8 weeks of age, it was significantly delayed in alcohol and pair-fed animals. Almost all morphological parameters of bone were lower in the alcohol groups. No significant difference in serum calcium levels, osteocalcin, or growth hormone levels were found, and small difference in calciotropic hormone levels was found between groups. The results indicated that chronic alcohol consumption during the age of bone development reduces bone density and peak bone mass in both cortical and cancellous bone. The mechanism whereby this effect occurs is not fully understood, but, our results suggest that the negative impact of alcohol on growing bone is not due to the secondary effects of altered bone mineral regulating hormones.

Acute and chronic effects of melatonin as an anticonvulsant in male gerbils.

Melatonin, a hormone produced in the pineal gland and released into the general circulation on a diurnal basis, has been implicated in many behavioral processes, where it has been shown to have anxiolytic, sedative, and anticonvulsant effects. Male gerbils (Meriones unguiculatus) injected daily with melatonin (25 micrograms, s.c.) exhibited a reduced seizure response to pentylenetetrazol (PTZ, 60 mg/kg, s.c.). The present studies determined 1) whether melatonin's effect was related to the time of day that it was administered and 2) whether a single acute injection of melatonin at various doses could produce anticonvulsant activity. Gerbils provided with 13 weeks of daily melatonin injections (25 micrograms, s.c.) exhibited fewer convulsions after PTZ treatment irrespective of the time of day melatonin was injected. In addition, the melatonin-treated gerbils had lower mortality rates (1/12) than the untreated or vehicle-injected gerbils (5/12). On the other hand, single acute injections of melatonin (0.1-10 mg/kg, i.p.) produced no anticonvulsant activity. It appears that the anticonvulsant effects of melatonin occur only after the animals are chronically exposed to the indole. In addition, melatonin's anticonvulsant ability may utilize a different mechanism than those involved in its endocrine effects, since no diurnal difference in melatonin's anticonvulsant activity was observed.

The relationship of nutritional copper to the development of postmenopausal osteoporosis in rats.

Factors that influence tissue copper concentration include age, diet, hormones, and pregnancy. In this study we altered diet independently, hormone (estrogen) independently, and various combinations of diet and hormone in animals of the same age to study the effects of ovariectomy complicated with dietary copper deficiency; a deficiency that has been demonstrated to cause bone defects. Sprague-Dawley rats were placed on various combinations of copper deficient or enriched diets before and/or after ovariectomy to determine if copper deficiency aggravated osteoporosis and if return to a copper-adequate diet alleviated it. In this study, ovariectomy did induce an osteopenia that was characterized by decreased trabecular bone. This osteopenia was slightly more severe with copper deficiency, but was not necessarily alleviated by the return of normal copper levels to the diet.

Rotation of dissecting groups among different cadavers provides students of anatomy with increased opportunity to observe human variations.

One way to increase medical students' awareness of anatomical variability is the dissection of different cadavers throughout laboratory coursework. This report covers such a procedure successfully instituted in a human gross anatomy course.

The suprachiasmatic nucleus controls the circadian rhythm of heart rate via the sympathetic nervous system.

The mammalian suprachiasmatic nucleus (SCN) controls a wide variety of circadian behavioral and physiological processes. The specific motor output pathway(s) by which these diverse processes are controlled are unknown. The only established motor output of this system is the regulation of pineal melatonin synthesis via the sympathetic nervous system. It is therefore possible that other peripheral circadian rhythms are regulated similarly. To address this issue, body temperature (BT), general activity (GA), wheel-running activity (WR), and heart rate (HR) were recorded in laboratory rats, and the effects of SCN lesion (SCNX) and of pharmacological sympathectomy with guanethidine (GUAN) on these multiple circadian rhythms were determined. The SCNX abolished circadian patterns in all motor outputs, whereas sham animals showed robust rhythms in all measures. In contrast, guanethidine, which depleted peripheral but not central catecholamine content, selectively reduced HR circadian rhythmicity. Other rhythms (BT, GA, and WR) were unaffected. These results suggest that the SCN influences some peripheral targets via circadian regulation of the sympathetic nervous system, and other circadian outputs are regulated via different, unknown pathways.

Temporary anestrus induced by injection of luteinizing hormone-releasing hormone in hamsters.

The photoperiod-induced anovulatory state of the Syrian hamster (Mesocricetus auratus) is characterized by a daily rhythm of circulating LH and FSH with a surge in blood levels occurring each afternoon. The association between anestrus and the presence of daily gonadotropin surges suggests that initiation of the characteristic daily rhythm of LH and/or FSH after transfer to short days results in blockade of ovulation. We tested whether daily injection of LHRH affects the 4-day rhythm of vaginal estrous cycles, induces daily surges of LH and FSH, blocks ovulation, and induces regression of the ovaries. LHRH (1 microgram/animal) was injected (s.c.) at 1630 h each day into females with regular 4-day estrous cycles. Injections began on the afternoon of estrus and continued for 12 days (experiment 1) or 8 days (experiments 2 and 3). In saline-injected controls, vaginal discharges occurred regularly at 4-day intervals during the injection period in all experiments. In contrast, vaginal estrus was not observed during the period of LHRH injection in any animals. Ova were found in oviducts of all saline-injected controls (8.9 +/- 0.8; n = 8) on the morning of estrus (after 8 injections), but no ova were found in oviducts of LHRH-injected females. Levels of plasma LH but not FSH increased after injection of LHRH each day, whereas the 4-day estrous cycle rhythm of gonadotropins was not disturbed in saline-injected controls. Vaginal estrous cycles and a 4-day rhythm of LH release resumed within 7 days after LHRH injections were stopped.(ABSTRACT TRUNCATED AT 250 WORDS)

Novel anticonvulsant action of chronic melatonin in gerbils.

Melatonin, a hormone from the pineal gland, was tested for its anticonvulsant effects in male gerbils. Daily administration of melatonin (25 micrograms injection-1, s.c.) for ten weeks reduced the number and severity of seizures (total convulsion score: 7.67 +/- 1.83 in controls vs 2.47 +/- 0.90 in melatonin-injected animals, p < 0.05) associated with the injection of the convulsant, pentylenetetrazol (PTZ, 60 mg kg-1, s.c.). However, neither 12 weeks of short photoperiod exposure (LD 10:14) nor biweekly administration of melatonin pellets altered PTZ-induced convulsions. Overall, melatonin-injected gerbils were better able to survive and respond to seizures than control animals. No melatonin-injected gerbils died during seizure induction (0/31) while 5 out of 33 control gerbils died after PTZ injection. The mechanism for melatonin's anticonvulsant effects could be due to a direct inhibitory action on neural activity or a conversion of melatonin to an anticonvulsant compound that resembles the kynurenines.

gamma-Aminobutyric acid, catecholamine and indoleamine determinations from the same brain region by high-performance liquid chromatography with electrochemical detection.

A new procedure for the measurement of gamma-aminobutyric acid, norepinephrine, dopamine, serotonin and 5-hydroxyindoleacetic acid from the same brain region was developed. In general, two separate high-performance liquid chromatographic runs were performed, one for the gamma-aminobutyric acid determination and one for the determination of the monoamines. The electrochemical detection of gamma-aminobutyric acid was determined by a new procedure that utilized a small aliquot of the brain sample prepared for monoamine measurement. This assay was linear and parallel between 6 and 200 ng per 20-microliters injection with 5-aminovaleric acid utilized as an internal standard. Inter-assay variability averaged 5% throughout the assay with gamma-aminobutyric acid values in the gerbil hypothalamus of 344 micrograms/g. The catecholamine assay has been characterized previously and utilizes 3,4-dihydroxybenzylamine as an internal standard with less than 5% variability. Norepinephrine, dopamine, serotonin and 5-hydroxyindoleacetic acid levels in the gerbil hypothalamus averaged 2922, 729, 797 and 272 ng/g, respectively. This new protocol allows a wide range of neurochemicals to be determined and evaluated from the same brain region.

Pineal serotonin is resistant to depletion by serotonergic neurotoxins in rats.

Administration of 3,4-methylenedioxymethamphetamine (MDMA) or para-chloroamphetamine (pCA) to adult rats is neurotoxic to serotonin (5HT) nerve terminals and cell bodies. MDMA (10 mg/kg) reduces 5HT levels in the frontal cortex, medial basal hypothalamus, and striatum acutely and 17 days after a series of multiple injections. The acute reductions occur within 1-2 hr after injection of doses greater than 3 mg/kg. A single injection of pCA reduces 5HT levels in the above mentioned brain regions as well as in the brain stem. However, none of these treatments are able to alter 5HT levels in the pineal gland. It appears that the pineal does not contain the 5HT reuptake system that is thought to be necessary for the neurotoxicity of MDMA and pCA.

Noradrenergic involvement in pinealectomy induced convulsions in gerbils.

Gerbils treated with various noradrenergic compounds were pinealectomized (PINX) or sham-operated (SHAM) and observed for convulsive activity. Although declines in cortical and hypothalamic norepinephrine content were found after alpha-methylparatyrosine (alpha MPT) or PINX, treatment with alpha MPT did not produce convulsions in SHAM gerbils or in gerbils which were PINX one week earlier. Likewise, alpha MPT did not increase the number or severity of convulsions observed in acutely PINX gerbils. Treatment with propranolol or isoproterenol did not have any effect on convulsive activity of PINX gerbils, except that isoproterenol increased the excitability of all the gerbils. In conclusion, reducing catecholamine content or modifying beta-noradrenergic receptor activity was not able to alter the convulsive activity which occurs after acute PINX.

Propranolol blockade of short photoperiod-induced gonadal regression: modification by melatonin injections or implants.

Male Syrian hamsters were exposed to short photoperiods (10L:14D) for ten weeks and received daily ethanolic saline (1:10), propranolol (125 micrograms, sc) or melatonin (25 micrograms, sc) injections at 16:45. Some hamsters also received melatonin implants (1 mg in 24 mg beeswax, sc) every two weeks in addition to the above treatments. Testicular weights were determined at the beginning, every three weeks, and at the end of the experiment. Serum hormone levels, hypothalamic amine levels and cortical amine levels were also determined at the conclusion of the experiment. Propranolol prevented the decline in testes weight produced by short photoperiod (P less than 0.001 versus control). Propranolol was not able to prevent daily injections of melatonin from producing gonadal regression. Melatonin implants blocked testicular regression produced by both short photoperiod and melatonin injections. No differences in hypothalamic or cortical amine levels were observed between the treatment groups. These results add further support to the suggestion that propranolol is acting at the pineal gland preventing melatonin synthesis and not at a post-pineal central site which would affect melatonin binding and action.

Effects of (+-)3,4-methylenedioxymethamphetamine (MDMA) on brain dopaminergic activity in rats.

Acute treatment with (+-)3,4-methylenedioxymethamphetamine (MDMA) at high doses (10 and 30 mg/kg, IP), but not lower doses increased locomotor activity in male rats. MDMA did not consistently produce any other stereotyped behaviors at any dose. Dopamine (DA) turnover rate as estimated by the ratio of brain tissue levels of 3,4-dihydroxyphenylacetic acid (DOPAC) over DA was decreased in the striatum for up to two hours after acute treatment with 10 mg/kg of MDMA. DA turnover rate was inconsistently decreased in the olfactory tubercle and medial basal hypothalamus, and was unchanged in the medial prefrontal cortex and the substantia nigra/ventral tegmental area. Two hours after a 30 mg/kg injection of MDMA, DA turnover rate was decreased in all brain areas tested. MDMA and d-amphetamine partially reversed a haloperidol-induced elevation of striatal DOPAC levels. In contrast, the nonamphetamine stimulant, amfonelic acid, enhanced haloperidol's effect. In chloral hydrate-anesthesized rats, MDMA injected IV partially inhibited spontaneous firing rate of DA neurons in the substantia nigra (34% decrease at 4 mg/kg of MDMA). Seventeen days after subchronic MDMA treatment (10 or 20 mg/kg, IP, twice per day for four days), DA and DOPAC levels were unchanged in all brain areas tested as compared to levels in control rats. It is concluded that acute treatment with high but not low doses of MDMA has a weak amphetamine-like effect on nigrostriatal as well as mesolimbic/mesocortical and tuberoinfundibular DA neurons in rats. Repeated treatment with MDMA does not appear to be toxic to mesotelencephalic or tuberoinfundibular DA neurons.

Transection of the pineal stalk produces convulsions in male Mongolian gerbils (Meriones unguiculatus).

The pineal gland has been implicated in the regulation of epileptic-like seizures in gerbils. Pinealectomy of gerbils results in seizures which affect approximately 80% of the gerbils. Telencephalic catecholamine levels are depressed by pinealectomy in gerbils and it has been suggested that the reduced norepinephrine (NE) levels are responsible for initiating or facilitating these seizures. The present study found that pineal stalk transection produced convulsions in gerbils which were indistinguishable from pinealectomy-induced convulsions. Stalk transection also produced a depression (38%) in parietal cortex NE levels which was similar to a depression observed in pinealectomized gerbils (52%). These depressions in cortical NE levels were not observed in other brain regions, specifically the hippocampus or the amygdala. However, a decline (25%) in hypothalamic NE levels was observed after stalk transection. These results suggest that the signals responsible for pinealectomy-induced convulsions are carried by the pineal stalk and that pinealectomy produces regional, not global, reductions in NE content.