Research reportThe expression of Fos within the suprachiasmatic nucleus of the diurnal rodent Arvicanthis niloticus
Introduction
There is extensive evidence that the hypothalamic suprachiasmatic nucleus (SCN) is the anatomical substrate for the primary circadian pacemaker in mammals. Lesions of the SCN abolish a variety of circadian rhythms [39], and transplants of fetal SCN tissue into animals with SCN lesions can restore a number of these rhythms [26]. In addition, a variety of rhythms intrinsic to the SCN have been documented in several species. For example, both in vitro and in vivo rates of glucose metabolism are higher in the SCN during the day than at night 40, 41, as are rates of single and multiple unit activity 14, 19, 20, 44. Thus, the SCN generates its own rhythms and is responsible for many circadian rhythms found in mammals.
Within the SCN of rats and hamsters the ventrolateral and dorsomedial regions are anatomically and functionally distinct. The ventrolateral region receives projections from the geniculohypothalamic tract (GHT) and the retinohypothalamic tract (RHT), both of which transmit visual input to the SCN 5, 15, 30. At least some of the fibers of the GHT contain neuropeptide-Y (NPY; 5, 16). The ventrolateral SCN is populated by cells containing VIP 27, 48, whereas cells in the dorsomedial SCN contain AVP [47]. These different areas are also characterized by different patterns of rhythmicity with respect to the expression and release of these various peptides. In the ventrolateral SCN the concentration of NPY is high around the transitions between light and dark phases of a 24-h light/dark (LD) cycle [21], and the peptide VIP as well as its messenger RNA peak during the dark phase of a 24-h LD cycle 2, 33, 45, 49. In contrast, the in vivo expression of AVP messenger RNA is higher during the day than at night 49, 50as is the release of AVP from the dorsomedial SCN in vitro 8, 13.
Rhythms in the expression of the nuclear phosphoprotein Fos have also been documented within the SCN of nocturnal rodents kept in various lighting conditions 3, 7, 10, 18, 24, 38, 43. The proto-oncogene c-fos encodes for the nuclear phospho-protein Fos, which binds to the DNA as a heterodimer with Jun and is involved in the regulation of gene transcription. The SCN of rats housed in a 12:12-h LD cycle exhibits a 24-h rhythm of Fos-IR which peaks during the light phase 9, 23. When different sub-regions of the rat SCN are examined separately, it becomes apparent that Fos-IR in the ventrolateral SCN is much higher during the light phase than the dark phase, whereas Fos-IR in the dorsomedial SCN is somewhat higher during the dark compared to the light phase [9]. In mice kept on a 12:12-h LD cycle, Fos-IR in the SCN is high immediately after lights on, drops by the middle of the day, and remains low until the lights are turned on again [7]. In rats kept in constant light (LL), Fos-IR is higher in the ventrolateral SCN during the subjective night than during the subjective day, but it remains unclear whether there is a rhythm in Fos-IR in the dorsomedial SCN 9, 10. In the SCN of mice and rats housed in constant dark (DD), no rhythm of Fos-IR is seen 7, 9, whereas hamsters kept in DD exhibit a slight rhythm in Fos-IR in the rostral SCN, with a peak in the middle of the subjective day [6]. In nocturnal rodents housed in DD, light pulses during the subjective night induce a dramatic increase in Fos-IR, whereas light pulses during the subjective day do not 38, 43. Thus, Fos-IR in the SCN of nocturnal rodents is expressed rhythmically in animals kept in LD cycles and LL, and responds to light pulses during DD in a time-dependent manner.
Research on neural mechanisms underlying mammalian circadian rhythms has focused primarily on nocturnal rodents, and nothing is currently known about how these mechanisms differ in diurnal and nocturnal species. Theoretically the differences could be due to differences within some subpopulation of SCN neurons, to differences in responsiveness to SCN signals, or to some combination of these two factors. With respect to three variables, rhythms within the SCN are similar in diurnal and nocturnal species. Rates of glucose utilization and electrical activity are higher during the day than at night in the SCN of both nocturnal and diurnal mammals 13, 19, 25, 37, 42, and immunoreactive AVP within the SCN is elevated during the day compared to the night in both humans and nocturnal rodents [17]. In spite of these findings, it remains possible that some aspect of SCN function differs in nocturnal and diurnal species, and is responsible for their differing patterns of rhythmicity.
Here we present two additional tests of the hypothesis that some aspect of SCN function differs in nocturnal and diurnal species. The diurnal animal model we have used is Arvicanthis niloticus, a small murid rodent found in sub-Saharan Africa. This species is diurnal with respect to its patterns of wheel-running, general activity, body temperature and copulatory behavior 22, 28. In the first study, we documented the rhythm in Fos-IR within the SCN of members of this species sacrificed at different phases of a 12:12-h LD cycle. In the second study we examined patterns of double-labeling of Fos and two different peptides found within SCN neurons, AVP and GRP.
Section snippets
Materials and methods
Animals used in this study were adult A. niloticus bred in the laboratory from a wild group captured in 1993 in Kenya. All animals were housed in plexiglass cages (38×34×16 cm) with same-sex siblings until 24–72 h before sacrifice, at which time they were individually housed. Animals were provided rodent chow (Harlan 8640 Teklad 22/5 Rodent diet) and water ad libitum, and were kept on a 12:12-h LD schedule; a red light (<5 lux) was kept on constantly.
Experiment 1: single labeling
Fos-IR was clearly observable within the nuclei of cells in the SCN, and the number of Fos-IR cells in the SCN changed significantly as a function of time of day (Fig. 1 and Fig. 2; F=18.792, P<0.001). The number of Fos-IR cells was highest 1 h after lights-on (ZT 1) and decreased progressively over the next 23 h (Fig. 1). Cell numbers at ZT 1 were significantly higher than at any other time point (Fisher's t-test, P<0.002), and those at ZT 5 and ZT 9 were significantly higher than at ZT 13 (P
Discussion
In the first experiment the concentration of Fos-IR cells in the SCN of A. niloticus was higher during the light hours than during the dark hours, and the transition from ZT 21 to ZT 1 was marked by a dramatic increase in Fos-IR. The number of Fos-IR cells in the SCN was highest at ZT 1, lower at ZT 5 and ZT 9, and still lower during the dark phase of a 12:12 LD cycle (ZT 13, 17, 21). A slight decrease in the levels of Fos-IR occurred over the course of the dark period, but this was not
Acknowledgements
We are grateful to Antonio Nunez, Cheryl Sisk and Kay Holekamp for comments on an earlier draft of the manuscript. This work was supported by NIMH Grant RO1 MH534/NS534433-01, and by the Neuroscience Training Program at Michigan State University.
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