Acute hypoxia-induced alterations of calbindin-D28k 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-binding proteins (CaBPs). Calbindin-D 28k is an EF-hand CaBP, which is highly expressed 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 buffering capacity and resulting in increase of intracellular and intranuclear [Ca ++ ]. Cerebellar tissues from normoxic fetuses were compared to fetuses obtained 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 (Hx0h) or were subsequently allowed to recover for 24 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 alterations of calbindin-D 28k immunoreactivity are qualitatively similar at all time points and include a distinctive intranuclear localization in subpopulations of PCs. A similar trend is demonstrated by immunoblotting. Subpopulations of TUNEL+/calbindin-D 28k - PCs lacking morphologic features of apoptosis or necrosis are demonstrated in Hx24h and Hx72h fetuses. The present study demonstrates an abrogating effect of perinatal hypoxia on calbindin-D 28k immunoreactivity in cerebellar PCs. The perturbation of this Ca ++ buffer protein in hypoxia-induced neuronal injury may herald delayed cell death or degeneration.