Differences in colocalization between Fos and PHI, GRP, VIP and VP in neurons of the rat suprachiasmatic nucleus after a light stimulus during the phase delay versus the phase advance period of the night

Abstract

Two groups of four rats each received a 15-minute light stimulus during the first part of the night (ZT14) and the second part (ZT19), respectively. After 45-60 minutes, the animals were killed by perfusion fixation. Adjacent Vibratome sections through the suprachiasmatic nucleus (SCN) were double-immunostained for the presence of peptide histidine isoleucine (PHI), gastrin releasing peptide (GRP) or vasoactive intestinal peptide (VIP) with Fos by using fluorophore-conjugated secondary antibodies. A few sections were triple-immunostained for PHI, GRP or VIP with vasopressin (VP) and Fos. Sections were analyzed with a confocal laser scanning microscope. It turned out that the ZT19 light stimulus induced 4.2 times more nuclear profiles in the SCN immunoreactive for Fos than the light stimulus given at ZT14. The SCN of control animals did not show any Fos immunoreactivity. After the ZT14 light stimulus, approximately 33% of the Fos profiles showed colocalization with a perikaryal profile immunoreactive for PHI, GRP or VIP, whereas at ZT19, this percentage had doubled to approximately 65%. After the light stimulus at ZT14, the relatively low Fos induction was numerically and proportionally most prominent in the PHI-immunoreactive perikarya. As compared with ZT14, the increase of Fos after the ZT19 light stimulus was most pronounced in the GRP-immunoreactive perikarya (21x) followed by VIP (15x) and PHI (5x). This outcome suggests that at least three different cell groups characterized by, respectively, PHI alone, GRP, and VIP fully or partly colocalized with PHI, play a prominent role during light-induced phase shifts: the PHI neurons during light-induced phase delays, the GRP and VIP/(PHI) neurons during light-induced phase advances.

Neurons containing gastrin-releasing peptide and vasoactive intestinal polypeptide are involved in the reception of the photic signal in the suprachiasmatic nucleus of the Syrian hamster: an immunocytochemical ultrastructural study

“…The occurrence of excitatory and inhibitory retinal synapses could be responsible for the two populations of Fos-positive and Fos-negative nuclei. In the rat, the colocalization of Fos-ir nuclei was evident in numerous, though not all, GRP-ir and VIP-ir cell bodies (Daikoku et al 1992;Mikkelsen et al 1994;Romijn et al 1996) or in GRP-ir neurons only (Earnest et al 1993).…”

“…The coexistence of immunoreactivities to GRP, VIP, and peptide histidine isoleucine amide (PHI) in the ventral part of the SCN suggested by light microscopy in the rat (Okamura et al 1986) has been clearly demonstrated as a colocalization in some neurons by confocal microscopy (Romijn et al 1996). In the Syrian hamster, the cells containing both GRP and VIP constitute only a subset of the GRP and VIP populations.…”

“…In their quantitative confocal microscopy study on the colocalization of Fos in GRP-ir, VIP-ir, or PHI-ir cells, Romijn et al (1996) have reported that more GRP-ir cells express Fos than VIP-ir or PHI-ir cells when light stimulation is applied in the middle of the night. At the same CT, a similar pattern of Fos expression within GRP-ir and VIP-ir neurons has recently been described in an unspecified species (Speh and Moore 1996).…”

“…These data are in agreement with our results. GRP and VIP are thus suspected of playing a prominent role during light-induced phase advances (Romijn et al 1996). In the hamster SCN, both GRP-ir and VIP-ir neurons are involved in light processing, but the question remains as to whether Fos is expressed only in GRP-ir or VIP-ir cells and/or in cells costoring GRP and VIP.…”

“…Third, the daily cycle in GRP and VIP mRNA (Okamoto et al 1991;Zoeller et al 1992) and immunoreactivity , which disappears in constant darkness (Shinohara et al 1993), suggests the involvement of these two peptides in the transmission of photic information. Finally, the expression of the Fos protein following light stimulation during the subjective night has been observed in the nuclei of rat SCN cells immunoreactive to GRP and VIP (Daikoku et al 1992;Earnest et al 1993;Mikkelsen et al 1994;Romijn et al 1996).…”