Acute hypoxia-induced alterations of calbindin-D-28k immunoreactivity in cerebellar purkinje cells of the guinea pig fetus at term

Purkinje cells (PCs) are vulnerable to hypoxic/ischemic insults and rich in calcium and calcium-buffering/sequestering systems, including calcium-bind ing proteins (CaBPs). Calbindin-D-28k is an EF-hand CaBP, which is highly e xpressed in PCs where it acts primarily as a cellular Ca++ buffer Elevation of [Ca++] in the cytosol and nuclei of PCs is pivotal in hypoxic/ischemic cell death. We hypothesize that hypoxia results in decreased concentration, or availability of calbindin-D-28k in PCs, thereby decreasing their buffer ing capacity and resulting in increase of intracellular and intranuclear [C a++]. Cerebellar tissues from normoxic fetuses were compared to fetuses obt ained from term pregnant guinea pigs exposed to hypoxia [7% FiO(2)] for 60 min. The pregnant guinea pigs were either killed upon delivery immediately following hypoxia (HxOh) or were subsequently allowed to recover for 23 h ( Hx24h) or 72 h (Hx72h). Fetal brain hypoxia was documented biochemically by a decrease in brain tissue levels of ATP and phosphocreatine. Compared to normoxic fetuses. there is a predominantly somatodendritic loss or decrease of calbindin-D-28k, immunohistochemical staining in PCs of HxOh (p < 0.005 ). Hx24h (p < 0.05), and Hx72h (p < 0.005) fetuses. Hypoxia-induced alterat ions of calbindin-D-28k immunoreactivity are qualitatively similar at all t ime points and include a distinctive intranuclear localization in subpopula tions of PCs. A similar trend is demonstrated by immunoblotting. Subpopulat ions of TUNEL+/calbindin-D-28k- PCs lacking morphologic features of apoptos is or necrosis are demonstrated in Hx24h and Hx72h fetuses. The present stu dy demonstrates an abrogating effect of perinatal hypoxia on calbindin-D-28 k immunoreactivity in cerebellar PCs. The perturbation of this Ca++ buffer protein in hypoxia-induced neuronal injury may herald delayed cell death or degeneration.