Serotonergic inhibition of light-induced fos protein expression and extracellular glutamate in the suprachiasmatic nuclei.

The present experiments were undertaken to explore a role for serotonin (5-HT) in modulating photic signal transduction and extracellular glutamate (Glu) concentration in the suprachiasmatic nuclei (SCN) of the Syrian hamster. Pretreatment with an i.p. injection of the serotonergic, quipazine, caused a marked decrease in the number of SCN cells expressing Fos protein-like immunoreactivity (Fos-LI) induced by a light pulse delivered during the latter part of the dark phase (7 h after lights-off; 55.6 +/- 7.5% of vehicle controls, P < 0.004). This effect of quipazine was dose-dependent and was limited principally to the ventrolateral region of the SCN. In a likewise manner, intra-SCN microinjection of quipazine inhibited light-induced Fos-LI in the ventrolateral SCN, indicating that the suppressive action of quipazine is centered in the SCN. In a separate experiment, localized perfusion of the SCN region with 5-HT using the microdialysis technique caused a significant reduction in the extracellular concentration of Glu. The effect was greater during the dark phase, compared to the light phase of the day-night cycle (60.7 +/- 6.8% vs. 39.3 +/- 6.8% maximal suppression, respectively; P < 0.05). Similar localized application of quipazine also decreased extracellular Glu (48.0 +/- 6.1% maximal suppression; P < 0.05). Collectively, these results are evidence for a serotonergic modulation of retinohypothalamic input in the SCN, which could involve a presynaptic inhibition of Glu release.

Differential timing of amino acid and 5-HIAA rhythms in suprachiasmatic hypothalamus.

In vivo brain microdialysis was used to characterize the daily pattern of extracellular excitatory amino acids (EAA; glutamate and aspartate), glutamine and 5-hydroxyindoleacetic acid (5-HIAA) in the region of the suprachiasmatic nucleus (SCN) in freely behaving male Syrian hamsters. Diurnal variations in the extracellular concentrations of EAA and 5-HIAA observed under 14:10-h light-dark (LD) photoperiod were confirmed by cosinor analysis. Peak levels occurred during the night, with the mean acrophasis for 5-HI-AA concentration preceding that for EAA by 1-2 h. Release of EAA was stimulated by K(+)-induced depolarization in a Ca(2+)-dependent manner and was not affected by tetrodotoxin. In the absence of light cues (constant dim red light, < 0.4 lx; DD) there was a significant time-of-day effect in peak glutamate concentration (occurring during subjective night), but the rhythmic pattern of 5-HIAA output was lost. There was no apparent direct temporal linkage between peaks in extracellular EAA or 5-HIAA and bouts of wheel-running activity under LD or DD. These results are evidence that 1) the daily release pattern of extracellular glutamate, but not 5-HIAA, is circadian in nature; and 2) this rhythm in glutamate is not based on Na+ channel-dependent action potentials. Finally, a discordance in the timing of peak nocturnal extracellular 5-HIAA and EAA concentrations was evident, possibly reflecting an interplay between serotonergic and EAA pathways in the SCN.

Suprachiasmatic nucleus neurochemistry in the conscious brain: correlation with circadian activity rhythms.

The aim of the research reported here was to provide information on the neurochemical processes that underlie the generation and entrainment of mammalian circadian rhythms. The studies were centered principally around the in vivo brain microdialysis technique for assessing the daily pattern of neurotransmitter activity in the suprachiasmatic hypothalamus of freely behaving Syrian and Siberian hamsters. This approach yielded several findings related to the activities of serotonergic and excitatory amino acid systems in the region of the suprachiasmatic nuclei (SNC). Specifically, we found that (1) there were daily variations in the extracellular concentrations of 5-hydroxyindoleacetic acid (5-HIAA) and glutamate (GLU) in the SCN region (highest levels of 5-HIAA occurred soon after lights-off, whereas GLU peaked later in the dark phase); (2) the daily rhythm of GLU, but not serotonin, in the SCN region appeared to be circadian in nature and was not driven by an external influence; (3) the rhythm in GLU measured in SCN microdialysate involved a tetrodotoxin-insensitive mechanism and did not appear to be directly linked to the expression of locomotor behavior; and (4) direct application of serotonin receptor agonists via the microdialysis probe suppressed the concentration of extracellular GLU in the SCN region, suggesting that serotonin may modulate GLU release in the SCN.

Diurnal variation in 5-hydroxyindole-acetic acid output in the suprachiasmatic region of the Siberian hamster assessed by in vivo microdialysis: evidence for nocturnal activation of serotonin release.

In vivo brain microdialysis was used to characterize the daily pattern of 5-hydroxyindole-acetic acid (5-HIAA) release in the region of the suprachiasmatic nuclei (SCN) in freely behaving male Siberian hamsters housed under 16L:8D. A marked diurnal variation in the concentration of extracellular 5-HIAA was apparent, with peak levels (147 +/- 5% of the daily mean; p < 0.05) occurring 2-3 h after lights-off. Smaller nocturnal rises in extracellular 5-HIAA were observed in the posterior hypothalamus and preoptic area (128 +/- 4 and 123 +/- 8% of the daily mean, respectively; both p < 0.05 vs. average daytime levels). Tryptophan loading increased 5-HIAA in SCN microdialysates by 44 +/- 6%, and this response was enhanced by localized perfusion with tetrodotoxin (TTX; 5 microM). Localized applications of KCl (150 mM) or veratridine (100 microM) decreased 5-HIAA by 62 +/- 5 or 49 +/- 11%, respectively. The effect of KCl was not significantly affected by specific calcium channel blockers. Perfusion with TTX markedly decreased SCN 5-HIAA during the dark phase, but had little effect during the light phase (42 +/- 8 vs. 12 +/- 5% suppression, respectively; p < 0.01). Addition of serotonin (3 microM) to the perfusate significantly stimulated 5-HIAA output. This treatment increased the release of 5-HIAA more during the dark than during the light phase (61 +/- 8 vs. 25 +/- 5%, respectively; p < 0.01).(ABSTRACT TRUNCATED AT 250 WORDS)

Increased FSH levels precede short photoperiod-induced anestrus in intact and unilaterally ovariectomized LSH/SsLak hamsters.

Anestrus brought about after 2-4 weeks of short photoperiod (SP) exposure in LSH/SsLak hamsters is preceded by impaired follicular development. Since the latter is critically dependent on adequate FSH levels, this study tested the hypothesis that SP might alter baseline or compensatory FSH levels prior to the onset of anestrus. Regularly cycling females in 141:10 (LP) were transferred to SP (8L:16D). Between 0800 and 0900 h on days 20 through 24 of SP exposure, half of the animals in diestrus II were unilaterally ovariectomized (UO), and the remaining animals were sham-operated (Sham-UO). Seven hours after surgery, blood samples were taken via cardiac puncture. All animals were killed the following morning at 0900 h. Uterine weights were significantly reduced in SP-exposed hamsters, yet the compensatory increase in FSH following unilateral ovariectomy was not affected; a tendency for higher levels was noted. On proestrus, serum and pituitary FSH levels of SP-exposed Sham-UO animals were significantly elevated over similarly treated LP-exposed hamsters. Interestingly in animals with severe follicular impairment, the highest FSH levels correlated with the lowest uterine weights. The data suggest that SP exposure does not impair compensatory FSH release or the ability of the ovaries to respond to UO. SP-induced elevations in FSH levels may result from reduced follicular secretion and reduced inhibin and/or estrogen levels.

Comparison of the effects of short photoperiod exposure and melatonin treatment in ovariectomized LSH/SsLaK hamsters.

The object of this study was to compare the effects of short photoperiod (SP) and melatonin (MEL) treatment on the reproductive axis in ovariectomized LSH/SsLak hamsters. Animals acclimatized in long photoperiods (LP) (14L:10D) and showing regular estrous cycles were ovariectomized. Half of the operated hamsters received Silastic capsules containing 17-beta estradiol (E2). On the following day the animals were further subdivided into three groups: the animals in one group received daily afternoon injects of melatonin (MEL), those in a second group were given the vehicle, and animals in the third group were transferred from LP to SP (8L:16D). All animals were killed after 30 days. In hamsters without E2 replacement, MEL or SP exposure significantly suppressed serum and pituitary FSH levels, although MEL was more effective in this regard. On the other hand, SP exposure did not change serum FSH levels in animals with E2 implants, whereas MEL effectively suppressed them. SP or MEL reduced serum LH levels to a similar extent in the absence of E2 replacement, yet in animals with E2 implants only MEL significantly lowered LH levels below LP E2-treated controls. This was in contrast to effects on the pituitary where both treatments were equally effective in the depression of LH content. Serum PRL levels were similarly suppressed by MEL or SP exposure in E2-treated hamsters. On the other hand, pituitary PRL levels were not affected by either treatment in animals with E2-containing capsules, whereas SP or MEL treatment both significantly depressed pituitary PRL contents in hamsters without E2 replacement. SP treatment lowered MBH LHRH contents in animals with E2-containing capsules; no other significant changes in hypothalamic LHRH were noted. The data suggest that daily treatment with 25 micrograms of MEL is generally more effective in the suppression of gonadotropin levels than SP exposure. It is suspected that the mode of administration of MEL, and its quantity, may interact with estrogen differently than SP in the induction of physiological changes and regulation of the LHRH system.

Early short photoperiod effects in female LSH/SsLak Syrian hamsters.

Previous experiments showed that LSH/SsLak female hamsters cease ovarian cyclicity after 2-4 weeks of short photoperiod (SP) treatment. This study was designed to reveal early hormonal and histological changes on day 1 of the estrous cycle (estrus) in SP-treated animals and changes brought about by the SP-induced acyclic condition. Groups of 8-10 animals were killed on day 1 after 0, 4, 8, 12, and 16 days of SP treatment; sera and pituitaries were saved for hormone determinations, and ovaries were prepared for histological analysis. Intraatrial cannulae were inserted into anovulatory animals, and blood samples were withdrawn on the first and second postoperative days; the animals were killed on the third postoperative day. PRL levels were significantly reduced in cycling animals after 16 days of SP exposure and diminished further in acyclic animals. Pituitary PRL did not change in cycling animals, although it was highly depressed in SP-treated acyclic animals. The estrous FSH surge, serum LH levels, and pituitary gonadotropin contents were not affected by SP in cycling animals; in anovulatory animals, pituitary gonadotropin contents were significantly increased, and daily afternoon gonadotropin surges were observed. Uterine weights were reduced in cycling animals and underwent a further reduction after cyclicity ceased. Ovarian analysis revealed that all cycling animals had the same number of recruited follicles, yet significantly fewer corpora lutea and an apparent increase in atretic antral follicles were observed after 16 days of SP exposure. These results suggest that SP-induced cessation of estrous cyclicity occurs abruptly. SP does not alter the secondary FSH surge, but might adversely affect maintenance of follicular growth and depress serum PRL levels.

Short photoperiod depresses castration response in female LSH/SsLaK hamsters.

This study was designed to examine the effects of short photoperiod (SP) exposure on gonadotropin and PRL levels in the presence and absence of estrogen treatment in ovariectomized LSH/SsLak hamsters. In experiment I, regularly cycling hamsters maintained in long photoperiod (14L:10D) were ovariectomized and Silastic capsules containing 2.0-mm columns of estradiol benzoate (EB) implanted simultaneously into half of the animals. On the following day, half of the animals in each treatment group were transferred to SP (8L:16D). After 20 days of SP or long photoperiod (LP) exposure, all animals were sacrificed by decapitation and their sera and pituitaries saved for hormonal determinations. The experimental protocol in experiment II was similar, except that two groups in each photoperiod received estrogen treatments; one group received 2.0-mm implants of 17-beta-estradiol (E2), whereas a second group received 10.0-mm E2 implants. SP treatment effected a reduction in serum LH and FSH levels in the absence of steroid replacement treatment. EB treatment depressed serum gonadotropin levels in LP animals, but did not alter levels in SP hamsters. In experiment II, LP- and SP-treated animals showed similar responses to E2 treatment, although different responses were noted in the two dosage groups. Pituitary gonadotropin contents became progressively decreased with increasing steroid levels and in certain groups showed SP-induced reductions. Serum and pituitary PRL levels increased in response to steroid treatment, but were not affected by SP treatment. In summary, 20 days of SP treatment caused gonad-independent reductions in gonadotropin levels and appeared to reduce the steroid feedback sensitivity of the hypothalamo-pituitary axis.(ABSTRACT TRUNCATED AT 250 WORDS)

Rapid cessation of estrous cyclicity and depressed castration response in short photoperiod-treated, inbred LSH/SsLaK hamsters.

This study examined the effects of transfer from long photoperiod (LP) to short photoperiod (SP) on the cessation of ovarian cyclicity and the castration response in inbred LSH/SsLak golden Syrian hamsters. Forty-six 8 to 10-wk-old female hamsters were acclimatized in LP (14L:10D; lights on at 0600 h) during which time animals were monitored for regular ovarian cyclicity. Twenty-six animals were transferred to SP (8L:16D; lights on at 0600 h) and examined daily for vaginal discharges. One day after the day of the first missed ovulation, individual SP-exposed animals were bilaterally ovariectomized; concomitantly, an LP control animal in diestrus I underwent the same procedure. Thirty days after ovariectomy, the hamsters were fitted with intra-atrial silastic cannulae. On the following two postoperative days, 0.6 ml blood samples were collected at 0700, 1200, 1400, and 1600 h for SP animals and at 0700, 1400, 1600 and 1800 h for LP controls. On the third day, the animals were decapitated and sera and pituitaries saved for determination of luteinizing hormone (LH), follicle-stimulating hormone (FSH) and prolactin (PRL) by radioimmunoassay (RIA). All SP-exposed animals displayed their last estrous discharge 14-34 days after transfer to SP (mean = 23.0 +/- 0.8 days). Their ovaries were characterized by the absence of corpora lutea, the presence of large atretic antral follicles, few growing follicles, and interstitium that was stimulated to varying degrees. Total and adjusted pituitary weights were decreased by SP exposure (p less than 0.001).(ABSTRACT TRUNCATED AT 250 WORDS)