Keiko Nakahara

Effect of lateral cerebroventricular injection of the appetite-stimulating neuropeptide, orexin and neuropeptide Y, on the various behavioral activities of rats

The effect of lateral cerebroventricular injection of the appetite-stimulating neuropeptide, orexin and neuropeptide Y (NPY), on the behavior of rats was investigated. An immediate increase in face washing activity was observed after injection of orexin A or orexin B, but not NPY. Orexin A had a more potent effect on face washing behavior than orexin B. Grooming and burrowing activities also increased significantly after injection of orexin A, whereas, orexin B significantly increased burrowing and searching behavior. Feeding behavior and food consumption increased dramatically within 10 min of injection of NPY. Although the significant increase in feeding behavior was also observed after injection of orexin A, total food intake did not change significantly. These results suggest that orexin may be involved in the regulation of several other behavioral activities in rats, besides feeding.

Effect of melatonin on circadian rhythm, locomotor activity and body temperature in the intact house sparrow, Japanese quail and owl

We compared the effect of melatonin on circadian rhythm, body temperature, and locomotion in the intact house sparrow, Japanese quail and owl. Daily treatment with melatonin at a fixed time did not entrain the free-running rhythm of locomotor activity in the house sparrow and the disrupted rhythm in Japanese quail under constant dim light. However, melatonin clearly inhibited movement for several hours after treatment. The duration of resting after injection of melatonin was dose-dependent. Body temperature was significantly decreased after melatonin treatment, the effect being more potent during the active phase than in the resting phase. Although this effect of melatonin on body temperature was also dose-dependent, the magnitude of the decrease in body temperature after injection of melatonin was greater in the house sparrow than in the Japanese quail. On the other hand, melatonin induced a further large decrease of body temperature in a nocturnal bird, the owl, whose pineal gland is degenerate. The decrease of body temperature was larger in the active phase than in the resting phase, and melatonin did not prevent movement in spite of the decrease in body temperature. These results suggest that the effects of melatonin on circadian rhythm, locomotor activity and body temperature differ among avian species, and that these mechanisms may not be linked to each other.

Individual pineal cells in chick possess photoreceptive, circadian clock and melatonin-synthesizing capacities in vitro

Chick pineal cells express a circadian rhythm of melatonin release under light-dark (LD) cycles, with an increase during the dark period and a decrease during the light period, and this rhythm persists under constant darkness (DD). We cultured individual single pineal cells with 15 microl of medium per well in a Terasaki plate and measured melatonin secretion every 12 h under LD, DL and DD. Individual cells secreted more melatonin during the dark period than during the light period under both LD and DL conditions, and those rhythmic secretions persisted under DD. These results suggest that individual pineal cells in chick have photoreceptive, circadian clock and melatonin-synthesizing capacities.

Effect of chronic treatments with ghrelin on milk secretion in lactating rats.

Ghrelin, a novel acylated peptide and endogenous ligand for growth hormone (GH) secretagogue receptor, was originally isolated from rat and human stomachs. In addition to its GH-releasing activity, ghrelin plays an important role in many physiological functions, including food intake, gastric acid secretion, neonatal development, and so on. In this study, the effect of daily treatment with ghrelin on milk production was investigated in lactating rats and the development of the pups was monitored. Daily subcutaneous injection of ghrelin into nursing dams for 8 days from parturition caused a significant increase in milk yield and litter weight gain. When litters nursed by ghrelin-treated and saline-treated dams were interchanged on day 4 of lactation, the growth curves were reversed. Daily injections of ghrelin also increased plasma GH levels. Northern blot analysis revealed that daily injection of ghrelin significantly increased mammary casein mRNA expression. In addition, RT-PCR analysis showed that a ghrelin receptor was present in the mammary glands of lactating rats. These results suggest that ghrelin may play an important role in milk production in lactating dams.

Effect of glucagon-like peptide-1 and -2 on regulation of food intake, body temperature and locomotor activity in the Japanese quail

To investigate the physiological roles of glucagon-like peptide-1 (GLP-1) and glucagon-like peptide-2 (GLP-2) in avian species, we elucidated the effect of intraperitoneal (i.p.) and intracerebroventricular (i.c.v.) administration of GLP-1 and GLP-2 on food intake, body temperature and gross locomotor activity in adult Japanese quail. Both i.p. and i.c.v. administration of GLP-1 suppressed food intake at 2, 4 and 12h after administration. Moreover, both i.p. and i.c.v. administration of GLP-1 significantly decreased both body temperature and gross locomotor activity 2h after administration. On the other hand, both i.p. and i.c.v. administration of GLP-2 had no effect on food intake, body temperature or gross locomotor activity. These results suggest that GLP-1 may have an important role in the regulation of food intake, body temperature and locomotor activity while GLP-2 may have no apparent effect on feeding regulation in adult Japanese quail.

Effects of microinjection of melatonin into various brain regions of Japanese quail on locomotor activity and body temperature

We previously reported that peripheral administration of melatonin resulted in simultaneous decreases in locomotor activity and body temperature in the Japanese quail. To examine the site of these melatonin-induced effects, we performed microinjection of melatonin into various brain regions of the Japanese quail. On the basis of the injection site the effects of melatonin were dissociated into four groups: decrease in only body temperature, decrease in only locomotor activity, decrease in both body temperature and locomotor activity, and a change in neither. The decrease in only body temperature was observed when melatonin was injected into areas of the diencephalon including the thalamus and hypothalamus, and the decrease in only locomotor activity was observed when melatonin was injected close to the nuclei septalis medialis and septalis lateralis. These results suggest that melatonin-induced decreases in body temperature and locomotor activity occur by melatonin acting at discrete sites in the brain.

Effect of neuromedin S on feeding regulation in the Japanese quail

Neuromedin S (NMS) was recently isolated from the brains of humans, mice and rats as an endogenous ligand for the orphan G protein-coupled receptors FM-3 and FM-4, which have been identified as neuromedin U (NMU) receptors 1 and 2, respectively. To investigate the role of NMS in avian species, we elucidated the effect of intracerebroventricular (i.c.v.) administration of rat NMS on food intake, body weight, body temperature and gross locomotor activity in adult Japanese quails. NMS significantly decreased food intake (and consequently body weight) in a time-dependent manner during 12-h light period, but increased both body temperature and gross locomotor activity. On the other hand, i.c.v. injection of rat NMU showed the reverse effects of NMS in Japanese quail. These results suggest that NMS may play an important role in regulating food intake and sympathetic nerve activity in the Japanese quail.

Regulation of GH secretagogue receptor gene expression in the rat nodose ganglion

It has been shown that the ghrelin receptor, GH secretagogue receptor (GHS-R), is synthesized in neurons of the nodose ganglion and then transmitted to axon terminals, where it binds to ghrelin. The orexigenic signal of ghrelin secreted from the stomach is transmitted to the brain via the vagal afferent nerve. To explore the regulation of GHS-R synthesis in the nodose ganglion, we examined whether or not GHS-R type a mRNA expression shows circadian rhythm, and is affected by starvation, vagotomy, or i.v. administration of gastrointestinal peptides. Nodose ganglion GHS-R mRNA levels showed a diurnal rhythm, being high during periods of light and low during darkness. Although starvation tended to increase the level of GHS-R mRNA, a more significant increase was observed upon re-feeding. Vagotomy decreased the level of GHS-R mRNA significantly in comparison with animals that underwent a sham procedure. Cholecystokinin and gastrin increased the level of GHS-R mRNA after 2 h, but after 4 h, the level decreased. These results suggest that GHS-R synthesis in the nodose ganglion is regulated centrally and peripherally by neuronal and humoral information, and that these dynamic changes of GHS-R mRNA expression may be involved in the regulation of feeding by ghrelin.

Comparison of feeding suppression by the anorexigenic hormones neuromedin U and neuromedin S in rats

We compared the central mechanisms of feeding suppression by the anorexigenic hormones neuromedin U (NMU) and neuromedin S (NMS) in rats. I.c.v. injection of either NMU or NMS dose dependently decreased 3-h food intake during the first quarter of a dark period. Pretreatment involving i.c.v. injection of a specific anti-NMS IgG blocked the suppression of food intake induced by i.c.v.- and i.p.-injected leptin, but anti-NMU IgG elicited no blockade. Quantitative PCR analysis revealed that i.c.v. injection of NMU or NMS caused a dose-dependent increase in CRH and proopiomelanocortin mRNA expression in the paraventricular nucleus (PVN) and arcuate nucleus (Arc) respectively. In tissue cultures of the Arc, secretion of α-melanocyte-stimulating hormone was stimulated by NMU and NMS, with more potent stimulation by NMS. The time-course curves of the increase in neuronal firing rate in Arc slices in response to NMU and NMS showed almost the same pattern, with a peak 10-15 min after treatment, whereas the time-course curves for the PVN slices differed between NMU and NMS. These results suggest that NMS and NMU may share anorexigenic effects, depending on physiological conditions.

Pituitary adenylate cyclase-activating polypeptide (PACAP) is involved in melatonin release via the specific receptor PACAP-r1, but not in the circadian oscillator, in chick pineal cells.

Pituitary adenylate cyclase-activating polypeptide (PACAP) stimulates melatonin release from pineal cells and modulates glutamatergic regulation of the suprachiasmatic circadian clock in rodents. We investigated whether PACAP is involved in melatonin release and the circadian oscillation system in chick pineal cells, and if so, whether its effects are mediated by the PACAP-specific receptor (PACAP-r1) or the vasoactive intestinal polypeptide (VIP) receptor. Chick pineal cells were maintained for 4 days under a 12-h light/dark cycle, and thereafter in constant darkness. In the dose-range 10(-10) to 10(-6) M, PACAP increased melatonin release dose-dependently during the 12-h light period on day 3 of culture, and the degree of stimulation was greater than that produced by VIP. VIP receptor antagonists only slightly inhibited PACAP-stimulated melatonin release. Simultaneous addition of VIP and PACAP produced almost additive melatonin release. Under constant dark conditions, 6-h pulses of PACAP started at zeitgeber times (ZT) 15, 21, 3 and 9 h in separate groups of pineal cells did not cause any phase shift in their melatonin rhythm. In addition, PACAP did not affect the light-induced phase advance (ZT 15 h) and delay (ZT 9 h) in melatonin rhythms. The expression of mRNA for the PACAP-r1 (including its splicing variant with a hop cassette) was observed in chick pineal cells. These results suggest that PACAP participates in melatonin release, but not in the circadian oscillator system, via the specific receptor PACAP-r1 in chick pineal cells.