The depression in rat pineal melatonin production after saline injection at night may be elicited by corticosterone

A hind-leg subcutaneous saline injection into rats at night elicits a decrease in N-acetyltransferase (NAT) activity and melatonin content of the pineal gland. The decrement in pineal melatonin production after saline injection is prevented by adrenalectomy. The present studies were undertaken to determine what factor(s) from the adrenal gland cause(s) the drop in pineal melatonin production after saline injection at night. In the first study, groups of intact and adrenal-demedullated male rats were given a saline injection at 23.10 h (3 h, 10 min after lights off) and their pineals were collected 15 or 30 min later. Pineal NAT activity was depressed in both intact and adrenal-demedullated rats at 15 min postinjection as compared to their respective control animals. Pineal melatonin levels exhibited a drop in intact animals at 15 min and in adrenal-demedullated rats at 30 min. In a second study, hypophysectomy was found to prevent the drop in nocturnal pineal NAT activity and melatonin levels normally associated with a hind leg injection of saline. Finally, in a third experiment, groups of hypophysectomized rats were injected i.p. with corticosterone at 23.10 h and killed 10, 25 or 40 min postinjection. Corticosterone injection in hypophysectomized rats produced a response similar to that caused by saline injection in intact animals: NAT activity was depressed at 10 min and melatonin content was lowered at 25 min. These results suggest that the adrenal-mediated depression in melatonin synthesis after saline injection at night in rats may be elicited by an adrenal cortical hormone (corticosterone) and apparently does not involve the release of factors from the adrenal medulla.     

Increased pineal melatonin content coupled to restricted water availability in a Pavlovian conditioning paradigm in rats

Summary The objective of this study was to assess whether rat pineal melatonin content could be modified in a classical conditioning paradigm. In rats kept under light (200 lux) from 06.00 to 18.00h daily, the time of lights off was selected as the unconditioned stimulus (US). Restricted water availability (from 10 min before to 10 min after light-dark, LD, transition) was the conditioned stimulus (CS). The conditioned and unconditioned responses were measured as the changes in pineal melatonin levels 4 h after LD transition. In animals under regular lighting conditions, lights out at 18.00 h (the US) caused a 4.4–7.8-fold increase of pineal melatonin concentration 4 h after later, when compared to animals maintained under light for the 4 h-period. After a training period of 7 days of restricted water availability (the CS), significantly augmented pineal melatonin levels were found in rats that were exposed to water but were maintained under light for the 4 h period after expected LD transition. The control animals for this experiment, i.e., rats which had undergone the training period, were kept for 4 h under light after expected LD transition, and did not receive water at LD transition, exhibited very low pineal melatonin levels. The conditioned increase of pineal melatonin content attained lower values than those in rats exposed to normal lighting conditions. It also fulfilled the contingency criterion, that is, it caused at trial a significant elevation of pineal melatonin content only when water availability was applied from 10 min previously to LD transition during training, and not 20 min after LD transition. After a training period of 7 days, restricted water availability applied 4 h before lights off (at 14.00 h), caused an enhanced production of melatonin 4 h later, regardless of the animals being exposed either to a dark or to a light environment. The results indicate that pineal melatonin production can be manipulated in a classical conditioning paradigm, when an appropriate CS stimulus is used.     

Influence of light irradiance on hydroxyindole-O-methyltransferase activity, serotonin-N-acetyltransferase activity, and radioimmunoassayable melatonin levels in the pineal gland of the diurnally active Richardson's ground squirrel

When Richardson's ground squirrels were kept under light:dark cycles of 14:10 h there was no nocturnal rise in pineal hydroxyindole-O-methyltransferase (HIOMT) activity. Conversely, the 10 h dark period was associated with large nocturnal rises in both pineal serotonin-N-acetyltransferase (NAT) activity and radioimmunoassayable melatonin levels. The nighttime rises in pineal NAT and melatonin were not suppressed by the exposure of the animals to a light irradiance of 925 mu W/cm2 during the normal dark period. On the other hand, when the light irradiance was increased to 1850 mu W/cm2 the rise in pineal NAT activity was eliminated while the melatonin rise was greatly reduced. When ground squirrels were acutely exposed to a light irradiance of 1850 mu W/cm2 for 30 min beginning at 5.5 h after lights out, pineal NAT activity and melatonin levels were reduced to daytime values within 30 min. The half-time (t 1/2) for each constituent was less than 10 min. Exposure to a light irradiance of either 5 s or 5 min (beginning at 5.5 h into dark period) was equally as effective as 30 min light exposure in inhibiting pineal NAT activity and melatonin levels. When animals were returned to darkness after a 30 min exposure to a light irradiance of 1850 mu W/cm2 at night, both pineal NAT activity and melatonin levels were restored to high nighttime levels within 2 h of their return to darkness. The results indicate that the pineal gland of the wild-captured, diurnal Richardson's ground squirrel is 9000 X less sensitive to light at night than is the pineal gland of the laboratory raised, nocturnal Syrian hamster.     

Changes in Pineal Indoleamines in Rats after Single Melatonin Injections: Evidence for a Diurnal Sensitivity to Melatonin

We recently determined that melatonin stimulated serotonin (5-HT) secretion from rat pineal glands by increasing 5-HT release from the pinealocytes (μM melatonin concentrations) and by inhibiting 5-HT uptake in the pineal sympathetic nerve endings (mM melatonin concentrations). The present study investigated whether a single melatonin injection could alter the content of indoleamines in the rat pineal gland, as well as its possible dependence on the daytime of administration. Melatonin (150 μg/kg) was i.p. injected at 8 time points (11.00 h, 14.00 h, 17.00 h, 20.00 h, 23.00 h, 02.00 h, 05.00 h and 08.00 h) to rats kept in 12:12 h light:dark cycle (lights on at 07.00 h). Melatonin injections in the afternoon (17:00 h) and late in the nighttime (02.00 h and 05.00 h) decreased pineal 5-HT content 90 min later. The levels of 5-hydroxyindoleacetic acid (5-HIAA) were also decreased 90 min after the melatonin treatment at 14.00 h, 17.00 h and 02.00 h. The effect of melatonin on 5-HT content was a long-lasting effect (still evident after 180 min) only when injected at 02.00 h, whereas 5-HIAA levels were found to be decreased 180 min after melatonin treatment at 14.00 h and 23.00 h. No changes in these compounds were detected 240 min after melatonin treatment. Moreover, melatonin did not change 5-hydroxytryptophan levels at any of the daytime points studied. By contrast, 90 min after the injection of melatonin at 20.00 h, an increased content of pineal N-acetylserotonin was observed. This effect of melatonin could be mediated through a phase alteration of the pineal N-acetyltransferase activity rhythm by acting on the suprachiasmatic clock, althought a direct melatonin effect on the pineal rhythmic function cannot be excluded. The effects of the hormone on 5-HT and 5-HIAA contents agree with previous findings on the inhibitory effect of pharmacological doses of melatonin on pineal 5-HT uptake, which presumably would result in a decreased intraneuronal content of 5-HT and its acid metabolite. These data point to an acute regulatory action of exogenous melatonin on the pineal melatonin synthesis pathway which seems to be limited to two daytime phases: the afternoon-early evening period and the second half of the night.     

A Possible Role of the Pineal Gland in Acute Immobilization-Related Suppression of Naloxone-Induced LH Release in Ovariectomized Estrogen-Primed Rats

It has been recently reported that acute immobilization stress almost completely suppresses the luteinizing hormone (LH) release induced by naloxone, a μ-opioid antagonist, in ovariectomized estrogen-primed rats. The present study examined the possible involvement of the pineal gland in the acute immobilization-related suppression of the naloxone-induced LH release. An intraventricular (ICV) injection of 15 μg naloxone produced an abrupt increase in circulating LH concentrations in non-stressed rats. The naloxone-induced LH release was completely eliminated when tested 60 min after the end of a 30 min session of acute immobilization. The same stress conditions did not affect LH-releasing hormone (LHRH)-induced LH release, suggesting that the stress-related suppression of the naloxone-induced LH release was a suprapituitary event. In chronically-pinealectomized rats, but not in sham-pinealectomized rats, naloxone injected 60 min after the end of the stress session evoked a significant increase in serum LH concentrations. However, naloxone injected ICV during the acute immobilization did not elicit LH release in either pinealectomized or sham-operated rats. Under non-stressed conditions, the LH secretory response to naloxone was similar in pinealectomized and sham-operated animals. The same stress (30 min immobilization) significantly increased pineal melatonin content as well as plasma melatonin concentrations in rats bearing intact pineal glands, indicating that stress actually affected the pineal function. These results provide evidence for a role of the pineal in the suppression of the LH response to naloxone after stress, but not during stress.     

Twenty-four-hour pineal melatonin synthesis in the vasopressin-deficient Brattleboro rat

The diurnal time course of pineal melatonin synthesis was analyzed simultaneously in the arginine vasopressin (AVP)-deficient Brattleboro rat (BB), the Long-Evans (LE) and Sprague-Dawley (SD) rat by means of radioenzymatic determination of the rate-limiting enzyme serotonin-N-acetyltransferase (NAT) and the melatonin content over a period of 24 h. While all 3 strains displayed a distinct day—night rhythm of melatonin synthesis (low day-time, high night-time values), BB rats generally exhibited lower NAT values as compared to LE and SD rats, though reaching a significant difference at 02.00 h only. Twenty-four-hour melatonin content was characterized by distinct nocturnal maxima in LE and SD rats, while BB rats showed a plateau-like nocturnal time course. Electrophysiological and pharmacological findings in SD rats point to an inhibitory influence of AVP upon pineal melatonin synthesis. The lack of AVP obviously does not result in disinhibition of pineal melatonin synthesis but rather in a different time course of pineal melatonin content. This might either be due to strain differences or to yet unknown compensatory mechanisms in BB rats.     

Suppression of pineal melatonin in Peromyscus leucopus by different monochromatic wavelengths of visible and near-ultraviolet light (UV-A)

The purpose of this study was to examine the effects of monochromatic visible and near-ultraviolet radiation (UV-A) on pineal melatonin suppression in the white-footed mouse, Peromyscus leucopus. To this end, mice were entrained to a daily cycle 8 h of light and 16 h of darkness. During the night when pineal melatonin contents were high, mice were individually exposed for 5 min to specific wavelengths of monochromatic light (10 nm half-peak bandwidths). Control animals received the same handling conditions but no experimental exposure. Pineal glands were collected from animals 18 min after the 5 min experimental exposure and were later assayed for melatonin content. In groups of animals exposed to equal photon densities (2.64 × 10¹⁵photons/cm²) of either 320, 340, 360, 500, or 560 nm, mean pineal melatonin content was significantly suppressed as compared to the unexposed control animals. The 640 nm wavelength (red) at the same photon density did not suppress pineal melatonin. These experiments are the first to demonstrate light-induced suppression of pineal melatonin in Peromyscus leucopus. In addition, these data reveal a novel finding: the suppression of pineal melatonin content by ultraviolet wavelength as low as 320 and 340 nm.     

Neither the pituitary gland nor the sympathetic nervous system is responsible for eliciting the large drop in elevated rat pineal melatonin levels due to swimming

Summary Since the pineal gland is an end organ of the sympathetic nervous system, stress might increase the synthesis of its hormone, melatonin. The stress of a 10 min swim, which elicits a marked rise in circulating catecholamines, causes a dramatic depression of high pineal melatonin levels at night within 15 min after swimming onset. N-acetyltransferase (NAT) activity is unaffected by the treatment at 15 or 30 min after swimming onset. Within 90 min after initiation of a 15 min swim, high nighttime pineal melatonin levels are restored while NAT values remain elevated. The swimming-induced reduction in high pineal melatonin levels is not influenced by either hypophysectomy, superior cervical ganglionectomy, prazosin (₁-adrenergic receptor blocker) pretreatment, yohimbine (₂-adrenergic receptor blocker) pretreatment, or reserpine (amine depletor) pretreatment. These results indicate that neither hormones secreted from the pituitary gland nor catecholamines secreted from the sympathetic nerves are involved in eliciting the dramatic reduction in elevated pineal melatonin levels in the rat.     

Parathion (O,O-dimethyl-O-p-nitrophenyl phosphorothioate) induces pineal melatonin synthesis at night

The effects of parathion on male rat pineal N-acetyltransferase (NAT) activity, hydroxyindole-O-methyltransferase (HIOMT) activity and pineal and serum melatonin levels at the end of light period (2000 h) and at night (2300 h and 0100 h) were studied. Additionally, pineal levels of 5-hydroxytryptophan (5-HTP), serotonin (5-HT), and 5-hydroxyindole acetic acid (5-HIAA) were estimated. Parathion was administered intragastrically at total doses (over 6 days) of either 6.5 or 13 mg/kg. Control rats received vehicle (corn oil) only. During the study, the rats were exposed to light:dark cycles of 14:10 with light off at 2100 h. Pineal NAT activity was increased at 0100 h following parathion administration at both doses, but HIOMT activity was unaffected. Pineal and serum melatonin levels were increased at night (2300 h and 0100 h) after the 13 mg/kg dose of parathion while the lower dose increased pineal melatonin only at 0100 h. Also, both doses decreased 5-HTP at 2000 h while the lower dose increased it at 2300; 5-HT was significantly decreased at 2300 h and 5-HIAA levels were lower but only significantly so for the 13 mg/kg dose at 2000 h. The results indicate that parathion has significant effects on pineal melatonin synthesis by mechanisms which remain unknown.     

Pituitary-adrenal and thyroid effects on melatonin content of the rat pineal gland

Based on clinical findings of diminished nocturnal serum melatonin levels in affective illness, we hypothesized that alterations in the pituitary-adrenal or thyroid axes of the rat might alter the nocturnal rise of melatonin content of the pineal gland in that species. Two experiments were conducted to investigate these issues. In the first, rats were injected for nine days with adrenocorticotropic hormone (ACTH) or corticosterone, timed to accentuate and prolong the normal circadian corticosterone rise. Although both these treatments produced significant elevations of serum corticosterone, there was no difference in pineal melatonin content during the day or night from that measured in control rats. In the second experiment, hypothyroidism was induced in rats by thyroid-parathyroidectomy, and hyperthyroidism was produced by injection of triiodothyronine (T3) for nine days. Despite clear evidence of metabolic and endocrine effects of these thyroid manipulations, pineal melatonin content was not altered during the day or night. The nocturnal increase of melatonin may have been phase-advanced in the hypothyroid group, although the experiment was not designed to detect such a shift. There thus is no evidence from this study in the rat to suggest that diminished nocturnal melatonin production in affective illness might be due to associated alterations in the pituitary-adrenal or thyroid systems.     

Alterations in components of the pineal melatonin synthetic pathway by acute insulin stress in the rat and Syrian hamster

Acute insulin stress increased plasma catecholamine levels in both the Syrian hamster and albino rat within 3 h after an intraperitoneal injection of either 5 or 10 units of insulin. In the rat, this stress caused a concurrent increase in pineal serotonin N-acetyltransferase (NAT) activity and melatonin content with no observable change in hydroxyindole-O-methyltransferase (HIOMT) activity. In the hamster, on the other hand, acute insulin stress did not alter pineal NAT activity, but depressed both HIOMT activity and melatonin content up to 3 h after the stress. These results present further evidence that catecholamines do not control hamster pineal melatonin synthesis by the same mechanism as observed in the rat.     

The human pineal gland responds to stress-induced sympathetic activation in the second half of the dark phase: Preliminary evidence

Summary Previous reports by our group and by others showed that the human pineal gland is unresponsive to stress-induced systemic sympathetic activation either during the day or 3 hrs after the beginning of darkness. In the present study, we investigated whether a longer period of dark exposure is required to demonstrate a stimulatory effect of stress-induced sympathetic activation on the human pineal gland. For this purpose, plasma melatonin levels were measured in six healthy men (aged 25–34 yrs) both in resting condition and before and after a physical exercise performed between 02.40 and 03.00 h, 30 min after exposure to bright light (2000 lux). Light exposure lasted from 02.10 h up to 04.00 h. The exercise consisted in bicycling on a bicycle ergometer at 50% of the personal maximum work capacity (MWC) for 10 min, followed by another 10min of bicycling at 80% of the MWC. In the same subjects, plasma melatonin levels were measured also without exposure to light and with no exercise (control dark condition). The results showed that physical exercise, although inducing a rapid and short-term general sympathetic activation (as shown by significant changes in cardiovascular parameters) was able to increase light-depressed plasma melatonin levels only 5 hrs after the end of the stress (p < 0.0001, group X time interaction, two-way ANOVA with repeated measures). These findings suggest that the human pineal gland is responsive to systemic sympathetic activation induced by physical stress in the second half of the dark phase.     

Lithium and melatonin in pigmented eye rats: Effects of dose and time of day

Lithium has been suggested to exert sane of its theraputic effects by modifying the function of the retinal-hypothalamic pineal pathway that is essential for the chronobiology of an organism (Seggie et al., 1983). Previous work was done in Wistar rats, an Albino species which lacks the enzyme for synthesis of eye pigment. This pigment is important in the regulation of light cued rhythms. The present project investigated effects of lithium in a pigmented eye strain. Adult male Long Evans rats were maintained individually on a 12 hour light/12 hour dark schedule with free access to water and one of three diets: (1) normal laboratory chow; (2) a low lithium diet: lab chow supplemented with 30 mM/kg of lithium chloride and (3) a high lithium diet: lab chow supplemented with 50 mM/kg of lithium chloride. Body weight and water intake were measured after six weeks on the diets. In Experiment I, separate groups of rats were sacrificed by rapid decapitation every 4 hours in the light/dark cycle. In Experiment II, animals were sacrificed every 90 min. between 12:00 and 20:00 hours during the dark cycle. Blood and pineal glands were collected for lithium determination and assay of melatonin by RIA. In Experiment I, plasma lithium levels were 0.35 ± 0.01 and 0.57 ± 0.02 mmEq/1 for the low and high diets. Serum and pineal melatonin evidenced the expected diurnal rhythms. The diets had no effect on these parameters except at 14:00 hours. At this time, two hours after lights out, the high diet but not the low diet group evidenced significantly lower serum melatonin levels than the control group (F = 4.91, df 2, 10, p < .05 for means of 14.0 ± 3.9 pg/ml vs 34.7 ± 5.6 pg/ml). In Experiment II plasma lithium levels were 0.39 ± 0.01 and 0.62 ± 0.01 mEq/1 for the low and high diets. Pineal and serum melatonin rhythms were again as expected and the diets had no effect except at 13:30 hours. At this time pineal melatonin content was significantly reduced by both diets. Control values were 2100 ± 133 pg/pineal, low diet 1222 ± 299 pg/pineal and high diet 905 ± 251 pg/pineal. (Control vs low F = 6.89, df 1, 8, p < .05; control vs high F = 15.5 df 1, 9, p < .01). Body weight was significantly reduced by both diets. Water intake was increased by the high diet and not affected by the low diet. In view of the consistency of time for the present significant findings, it is suggested that lithium is exerting a subtle and short-lived effect on pineal melatonin levels during the period shortly after the switch to darkness and maybe influencing the characteristics of the “turn-on” effects of this important timing cue. In Albino rats, diet supplemented with 50 mM/Kg of lithium chloride affected the pattern of serum melatonin levels between 14:00 and 22:00 hours (Seggie et al., 1983). Thus, a strain effect of lithium cannot be ruled out.     

Studies on pineal melatonin levels in a diurnal species,the Eastern chipmunk (Tamias striatus): Effects of light at night,propranolol administration or superior cervical ganglionectomy

Summary Five experiments were carried out on the control of melatonin levels in the pineal gland of a diurnal species, the Eastern chipmunk (Tamias striatus). We confirmed that the exposure of chipmunks to fluorescent white light of 3,981–4,304 lux during the normal dark period does not prevent the rise in pineal melatonin levels normally associated with darkness. Also, the administration of propranolol (20mg/kg) at 8 p.m. did not block the rise in pineal melatonin in animals exposed to either dark or light at night. Similarly, if chipmunks received propranolol 4 hours into the dark phase, pineal melatonin levels were not depressed 2 hours later. When animals were superior cervical ganglionectomized, however, the pineal content of melatonin remained low regardless of whether the animals were exposed to darkness or light at night. The exposure of chipmunks acutely to light at midnight (4 hours after darkness onset) had only a slight depressive effect on pineal melatonin 30 min later; by comparison, when chipmunks were acutely exposed to light at 3 a.m. (7 hours after darkness onset) daytime pineal melatonin levels were reached within 15 min after light onset. These findings in a diurnal species, the Eastern chipmunk, differ markedly when compared to previously reported observations on nocturnal laboratory rodents.     

Pineal indoleamine metabolism in pyridoxine-deficient rats

Pyridoxine deficiency causes physiologically significant decrease in brain serotonin (5-HT) due to decreased decarboxylation of 5-hydroxytryptophan (5-HTP). We have examined the effect of pyridoxine deficiency on indoleamine metabolism in the pineal gland, a tissue with high indoleamine turnover. Adult male Sprague-Dawley rats were fed either a pyridoxine-supplemented or pyridoxine-deficient diet for 8 weeks. Pyridoxine deficiency did not alter the pattern of circadian rhythm of pineal 5-HT, 5-hydroxyindoleacetic acid (5-HIAA), N-acetylserotonin (NAS), and melatonin. However the levels of these compounds were significantly lower in the pineal glands of pyridoxine-deficient animals. Pineal 5-HTP levels were consistently higher in the pyridoxine-deficient animals and a conspicuous increase was noticed at 22.00 h. Increase in pineal NAS and melatonin levels caused by isoproterenol (5 mg/kg at 17.00 h) were significantly lower (P less than 0.05) in the pyridoxine-deficient animals. Treatment of pyridoxine-deficient rats with pyridoxine restored the levels of pineal 5-HT, 5-HIAA, NAS, and melatonin to values seen in pyridoxine-supplemented control animals. These results suggest that 5-HT availability could be an important factor in the regulation of the synthesis of pineal NAS and melatonin.     

Castration Reduces the Nocturnal Rise of Pineal Melatonin Levels in the Male Rat by Impairing its Noradrenergic Input

The effects of castration and testosterone treatment on pineal day-night rhythms were studied in male rats. Bilateral gonadectomy was performed at 21 days of age. Testosterone propionate was given subcutaneously to castrated animals in a dose of 10 μg/100 g body weight during two consecutive days before sacrifice. Animals were killed 40 days after gonadectomy at four different times of a 12:12 h light-dark cycle (1600, 2400, 0400 and 0800h). Tyrosine hydroxylase activity was measured in individual pineals by means of high-performance liquid chromatography determination of L-DOPA formed. Pineal levels of norepinephrine, dopamine, 5-hydroxytryptamine and 5-hydroxyindole acetic acid were determined by high-performance liquid chromatography with amperometric detection, while pineal melatonin content was measured by radioimmunoassay. Castration abolished the day-night rhythms of pineal tyrosine hydroxylase activity and norepinephrine content, both by elevating their daytime levels and by blocking their nocturnal rise. In addition, gonadectomy drastically modified pineal indoleamine metabolism by increasing daytime levels of both 5-hydroxytryptamine and 5-hydroxyindole acetic acid, and by reducing the nocturnal elevation of pineal melatonin content. Testosterone treatment was unable to prevent the effect of orchidectomy on pineal rhythms of tyrosine hydroxylase activity, 5-hydroxytryptamine or 5-hydroxyindole acetic acid content, however it partially restored the day-night pineal rhythms of both norepinephrine and melatonin content. These results are indicative of a possible participation of reproductive hormones in the control of pineal rhythmic activity in the male rat. Apparently, since gonadectomy abolished the nocturnal rise of both pineal tyrosine hydroxylase activity and norepinephrine content, the primary site of action of reproductive hormones could be at the level of the superior cervical ganglion.     

Food restriction retards aging of the pineal gland

The effects of chronic (40%) food restriction from 6 weeks of age were studied in aging male Fisher 344 rats. When compared with 3-month-old, ad libitum fed rats, pineal N-acetyltransferase (NAT) activity had declined to less than 30% and pineal and serum levels of melatonin to 40% after 28 months when feeding had been ad libitum. Food restriction significantly retarded this development (P less than 0.05) giving NAT and melatonin levels which were twice as high as in the ad libitum fed group. Nighttime levels of pineal serotonin (5-HT) were similar in food-restricted and ad libitum fed old rats but were nearly twice as high (P less than 0.05) as in young rats. There was also a tendency for increased production of 5-hydroxyindoleacetic acid (5-HIAA) in the pineal gland with higher levels of 5-HT. It is concluded that aging in the rat (Fisher 344) is accompanied by a reduction of pineal NAT activity, thereby reducing the production of melatonin and causing a buildup of 5-HT in the pineal gland. It is furthermore proposed that food restriction, which markedly increases the life span and reduces age-related physiological deterioration and diseases in many animals, may mediate some of its effects through a sustained pineal activity in old age.     

Peptidergic modulation of serotonin release from cultured rat pinealocytes

This paper describes the effects of beta-adrenergic and peptidergic inputs on serotonin (5-HT) synthesis, outflow and metabolism into melatonin in cultured dissociated rat pinealocytes. The spontaneous outflow of 5-HT from pinealocytes was high as demonstrated by the elevated levels of extracellular 5-HT accumulated in the medium (about 5 ng/h/70,000 pineal cells). The beta-adrenergic agonist isoproterenol (ISO) used at concentrations up to 10(-6) M induced a moderate (+20-40%) increase in intra- and extracellular 5-HT levels together with a large release of melatonin. At a higher ISO stimulation (10(-5) M), the intra- and extracellular levels of 5-HT were significantly (-25-30%) reduced whereas melatonin secretion was dramatically increased. This is interpreted as a large 5-HT mobilization for melatonin synthesis and release, consequently reducing both the intracellular pool and outflow of 5-HT. The peptides vasoactive intestinal peptide (VIP) and pituitary adenylate cyclase activating peptide (PACAP) up to 10(-7) M induced always a moderate (+20-30%) increase in intra- and extracellular levels of 5-HT. However, the use of nM concentrations of VIP or PACAP together with 10(-6) M ISO induced a decrease in 5-HT outflow (-25-30%) and a dramatic increase in melatonin secretion as did 10(-5) M ISO alone. Neuropeptide Y (NPY) is another pineal peptide which induced a stimulation of 5-HT outflow (+30-40%) although its effect on melatonin release was marginal. The above results are discussed in term of the multineuronal regulation of the synthetic and secretory activities of the rat pineal gland.     

Suprachiasmatic control of melatonin synthesis in rats: inhibitory and stimulatory mechanisms

The suprachiasmatic nucleus (SCN) controls the circadian rhythm of melatonin synthesis in the mammalian pineal gland by a multisynaptic pathway including, successively, preautonomic neurons of the paraventricular nucleus (PVN), sympathetic preganglionic neurons in the spinal cord and noradrenergic neurons of the superior cervical ganglion (SCG). In order to clarify the role of each of these structures in the generation of the melatonin synthesis rhythm, we first investigated the day- and night-time capacity of the rat pineal gland to produce melatonin after bilateral SCN lesions, PVN lesions or SCG removal, by measurements of arylalkylamine N-acetyltransferase (AA-NAT) gene expression and pineal melatonin content. In addition, we followed the endogenous 48 h-pattern of melatonin secretion in SCN-lesioned vs. intact rats, by microdialysis in the pineal gland. Corticosterone content was measured in the same dialysates to assess the SCN lesions effectiveness. All treatments completely eliminated the day/night difference in melatonin synthesis. In PVN-lesioned and ganglionectomised rats, AA-NAT levels and pineal melatonin content were low (i.e. 12% of night-time control levels) for both day- and night-time periods. In SCN-lesioned rats, AA-NAT levels were intermediate (i.e. 30% of night-time control levels) and the 48-h secretion of melatonin presented constant levels not exceeding 20% of night-time control levels. The present results show that ablation of the SCN not only removes an inhibitory input but also a stimulatory input to the melatonin rhythm generating system. Combination of inhibitory and stimulatory SCN outputs could be of a great interest for the mechanism of adaptation to day-length (i.e. adaptation to seasons).     

Effects of growing tumours on pineal melatonin levels in male rats

Summary The effect of transplantable tumours (Yoshida Sarcoma) on pineal melatonin content was studied in Wistar rats. A negative correlation between pineal melatonin content and size of growing tumours was observed. Effects of chronic treatment with drugs interfering with melatonin or serotonin biosynthesis on tumour growth as well as the influence of tumour growth on pineal melatonin content were investigated. Tumour growth enhancement observed after administration of some substances producing a decrease of pineal melatonin is discussed. Pinealectomy stimulates malignant growth. This may be caused by lack of endogenous melatonin.