MORPHOFUNCTIONAL HETEROGENEITY WITHIN THE RAT PINEAL GLAND: A LESSON FROM CREB/PCREB.

E.M. MUÑOZ; M.E. GUIDO; L.E. FARIAS ALTAMIRANO

Mendoza

Congreso; XXXIV Reunión Anual de la Sociedad de Biología de Cuyo; 2016

Sociedad de Biología de Cuyo (SBCuyo)

The circadian modulation of physiology and behavior facilitates adaptation to environmental changes. The pineal gland (PG) is one of the principal effectors and regulators of the mammalian circadian timing system by producing nocturnal melatonin. The PG is under sympathetic regulation via local norepinephrine (NE) release at night. In rat, phosphorylation of the transcription factor CREB (cAMP-responsive element (CRE)-binding protein) at serine 133 is essential to initiate the expression of the aanat gene. This gene encodes AANAT, one of the pivotal enzymes in the melatonin synthesis. To challenge the concept of pineal homogeneity, we analyzed CREB and pCREB at different ZTs (Zeitgeber time; L:D 12:12). We performed immunolabeling (IHC) and confocal microscopy in rat PG sections and Western blot (WB) analysis in whole PG protein extracts. The immunoblottings mostly confirmed previously published data for both protein forms. On the other hand, IHC revealed heterogeneity within the PG. While total CREB was found in pinealocyte nuclei at ZT6, 14 and 18, diversity in immunoreactive granule sizes and nuclear distribution patterns was seen throughout the L:D cycle. CREB present in the nuclei of interstitial cells showed a finer and more homogeneous distribution. For nocturnal pCREB, the signal varied from pinealocyte to pinealocyte with higher expression at ZT18. The phosphorylated form was negative in pinealocytes at ZT6. The presence of pCREB in interstitial cells at different ZTs infers that its phosphorylation may be independent of the nocturnal NE. These findings are of special interest because they suggest that melatonin synthesis varies significantly among individual pinealocytes. This cellular heterogeneity further implies that the pinealocyte population may have a greater need to synchronize its nocturnal melatonin release.