The microglia regulate pineal gland shape and function

M.P. IBAñEZ RODRíGUEZ; S. NOCTOR; E. MUÑOZ

Galveston, TX.

Congreso; GRC on Pineal Cell Biology; 2014

Gordon Research Conferences

In rat, the pineal gland (PG) begins as an evagination of the diencephalic roof at around E15 and becomes a solid organ at the end of the gestation period. In this study we characterized the major cellular and morphological events that take place in the developing PG from E15 to E21. Also, we studied the adult PG in normal conditions (Sham) and after disrupting the PG sympathetic innervation via bilateral superior cervical ganglionectomy (SCGx). In this case, PGs were collected at 3, 6, 9 and 13 days after surgery. We examined the expression patterns of the transcription factor Pax6; the astrocyte markers vimentin (vim), GFAP and S100â; the microglia markers Iba-1 and Isolectin IB4 which also labels blood vessels; the mitotic markers pSer10-histone H3 (pH3) and PCNA, and the neuron-specific class III â-tubulin (Tuj1). Pax6 was expressed in pinealocyte precursors throughout PG development, with the highest nuclear levels at early stages of embryogenesis. A few pH3-positive mitotic cells were present in the apical region of the stratified neuroepithelium of the pineal anlage from E15 to E17, but they were more abundant and randomly distributed in later stages. Interestingly, these mitotic cells showed basal Pax6 levels. Vim was expressed in the embryonic PG while GFAP and S100â were not detected. In the PG recess, Pax6/vim-positive cells were radially disposed. In later stages, especially at E18 and E19, cells immunolabeled for both Pax6 and vim were observed in rosette-like arrangements. At E20 and E21, high levels of vim expression were confined to a few randomly distributed cells. Iba-1/ IB4-positive microglial cells, likely derived from choroid plexus and meninges, colonized the PG from E15 onwards. They exhibited morphological features similar to those of active cells, and often they were observed in close proximity of Pax6- expressing precursors. To further understand the role of PG microglia, we performed bilateral SCGx to challenge the local microglial cells. The presence of degenerating nerve fibers activated PG microglia. Although these cells were observed in the entire SCGx gland, they were more abundant in the pineal stalk and proximal region, and they were located mainly close to IB4-positive blood vessels and in tight spatial relationship with degenerating Tuj1-positive nerve fibers. In contrast to the embryonic PG, GFAP and S100â were expressed in the adult PG in a region-dependent manner. A heterogeneous population of astrocytes that expressed one or both markers was observed in both Sham and SCGx conditions, being more abundant in denervated PGs. In the adult PG, a few cells were positive for Pax6; these cells were located perivascularly in close relationship with microglial cells, which in some cases acted as cellular bridges between those potential precursor cells and astrocytes. Interestingly the expression of Pax6 was slightly higher in the SCGx PGs. The cells positive for the mitotic marker PCNA were more abundant in the denervated glands and they were also immunoreactive for Iba-1. In brief, the PG evolves from Pax6/vim-positive precursor cells capable of adopting different morphologies and arrangements throughout development to give rise the main cell type, the pinealocyte, and a subpopulation of interstitial cells that is also present in the adult PG. The ganglionectomy activates the local microglia and maybe leads to the dedifferentiation of the PG. We speculate that microglia modulate PG morphogenesis and have a role in maintaining the adult phenotype by actively interacting with precursor cells, astrocytes, nerve fibers and blood vessels.