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Adverse effects of an active fragment of parathyroid hormone (PTH1–34), a blood Ca2+ level-regulating hormone, were examined using rat hippocampal slices in organotypic culture. Exposure of cultured slice preparations to 0.1 μM PTH1–34 for 60 min resulted in a gradual increase in the intracellular Ca2+ concentration ([Ca2+]i); this effect was most obvious in the apical dendritic region of CA1 subfield. When PTH1–34 at a lower concentration (1 nM) was added to the culture medium and its toxic effects examined using a propidium iodide intercalation method, significant toxicity was seen 3 days after exposure and increased with time. Cells in the CA1 region seemed more vulnerable to the hormone than cells in other regions. At 1 week of exposure, the toxic effects were dose-dependent over the range of 0.1 pM to 0.1 μM, the minimum effective dose being 10 pM. The adverse effects were not induced either by the inactive fragment, PTH39–84, or by an active fragment of PTH-related peptide (PTHrP1–34), an intrinsic ligand of the brain PTH receptor. The PTH1–34-induced adverse effects were significantly inhibited by co-administration of 10 μM nifedipine, an L-type Ca2+ channel blocker, but not by co-administration of blockers of the other types of Ca2+ channel. The present study demonstrates that sustained high levels of PTH in the brain might cause degeneration of specific brain regions due to Ca2+ overloading via activation of dihydropyridine-sensitive Ca2+ channels, and suggests that PTH may be a risk factor for senile dementia. Keywords: Parathyroid hormone, PTH1–34, PTHrP, hippocampal organotypic culture, intracellular Ca2+ concentration, L-type Ca2+ channel, nifedipine, propidium iodide, senile dementia Introduction Evidence demonstrating dysfunction of Ca2+ homeostasis during aging is accumulating, and a relationship with the incidence of dementia has been suggested (Fujita, 1990). It is now well established that an increase in the intracellular Ca2+ concentration [Ca2+]i leads to neurotoxicity (Choi, 1988; Ogura et al., 1988), and aging appears to be a major risk factor for Alzheimer's disease and several other neurodegenerative conditions, which suggests that an increase in [Ca2+]i could contribute to the increased vulnerability of aged neurons (Kipen et al., 1995; Satrustegui et al., 1996; Thibault & Landfield 1996; Porter et al., 1997). The regulation of blood Ca2+ levels in normal subjects results from well-designed Ca2+ homeostatic mechanisms that depend on specific hormones, such as parathyroid hormone (PTH) and calcitonin, which regulate the absorption of dietary Ca2+, reuptake of Ca2+ from the renal tube and Ca2+ retention in osteocytes. However, in elderly subjects, especially those with osteoporosis, and in Alzheimer patients, it has been shown that blood Ca2+ levels are lower and thus PTH levels higher, than in normal subjects (Desterhorft et al., 1994). In a previous study, we demonstrated that PTH causes a gradual increase in the [Ca2+]i by activating dihydropyridine-sensitive Ca2+ channels via specific PTH receptors in the brain (Hirasawa et al., 1998). This result suggested to us that high blood PTH levels might cause an increase in the [Ca2+]i in brain cells and thus induce degeneration of cells in the brain. Since the blood-brain-barrier in elderly subjects is reported to be less tight than in younger subjects (Ueno et al., 1997), the adverse effects of PTH may yield important clues in elucidating the causal relationship between the deterioration in Ca2+ homeostasis and the incidence of senile dementia. In the present study, we examined the adverse effects of PTH1–34, an active fragment of PTH, on organotypic hippocampal slice cultures using the propidium iodide (PI) intercalation method, and found that PTH1–34 had a marked toxic effect on the tissue due to dihydropyridine-sensitive Ca2+ channel activation. |
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