Search

In the present study, we tested whether the pineal organ of ayu (Plecoglossus altivelis), an osmerid teleost close relative of salmonids, harbours a circadian oscillator regulating rhythmic melatonin release using flow-through culture. The pineal organ maintained under light/dark cycles released melatonin in a rhythmic fashion with high levels during the dark phase. A circadian rhythm of melatonin release persisted in constant darkness for at least four cycles. Characteristics of the circadian rhythm (free-running period, phase and amplitude) exhibited small variations among cultures when the data was normalized, indicating that this system is sufficient for the analysis of the circadian rhythm both at qualitative and quantitative levels. Six-hour extension of the light phase from the normal onset time of the dark phase or exposure to constant light for 36 or 48 h before transfer to constant darkness significantly inhibited melatonin release. Phase shifts in the circadian rhythm of melatonin release were also observed. Thus, the ayu pineal organ contains all the three essential components of the circadian system (a circadian clock, the photoreceptor responsible for photic entrainment of the clock, and melatonin generating system as an output pathway). This system should provide a useful model for analysing the physiological and molecular basis of the vertebrate circadian system. In addition, further comparative studies using salmonids and related species including ayu will provide some insight into the evolution of the roles of the pineal organ in the vertebrate circadian system.

Nitric oxide and estrogen have been shown to play a critical role in the control of female reproductive function. In order to determine an anatomical relationship between nitric oxide generating neurons and estrogen target neurons, NADPH-diaphorase histochemistry was combined with estrogen receptor immunohistochemistry in the female medial preoptic area. While only a few weakly stained neurons for NADPH-diaphorase were found in ovariectomized control rats, a drastic increase in NADPH-diaphorase activity was observed in the medial preoptic nucleus of estradiol-treated ovariectomized animals. The total number of NADPH-diaphorase neurons in the estradiol-treated group increased three-fold relative to controls, and more than 80% of those neurons contained estrogen receptor-immunoreactivity in their nuclei. Since neuronal NADPH-diaphorase is nitric oxide synthase, the present result suggests that nitric oxide synthase activity can be positively regulated by estradiol in neurons containing estrogen receptor in the female medial preoptic nucleus.

Effects of gonadal steroids on numbers of neurons containing estrogen receptor (ER) and/or substance P (SP) were examined in the anteroventral periventricular nucleus (AVPV) of female and male rats by double-labeling immunohistochemistry employing antibodies specific for ER and SP. Animals were gonadectomized and received subcutaneously either oil alone (Control group), sequential injections of estradiol benzoate and oil (EB + Oil group), or those of EB and progesterone (EB + P group). In the female control rat, a large population of ER-immunoreactive (IR) cells were found clustered throughout the AVPV. They were counted more than 2,000 in total of 4 sections in this nucleus. On the contrary, SP-IR neurons were scarcely observed in the same area of this group. Administration of estrogen to female animals decreased the total number of ER-IR cells to 67% of the control group. In contrast to the supressive effect of estrogen to its own receptor, it induced SP-IR neurons in the AVPV of the female. Approximately 50-80 SP-IR neurons were counted in the 4 sections, and 59% of these neurons expressed ER-IR material in their nuclei. In the female EB + P group, the number of ER-IR neurons also decreased to 79% of the control group. Although the number of SP-IR neurons in this group decreased to 32% of that in the EB + Oil group, a ratio of coexistence of ER-IR material in these neurons increased to 75%. The male control group contained a smaller population of ER-IR cells relative to the female control (1497 vs 2143).(ABSTRACT TRUNCATED AT 250 WORDS)