Stimulation of calcium uptake in cultured rat hippocampal neurons by 2,3,7,8-tetrachlorodibenzo-p-dioxin
ABSTRACT This study examined the effects of 2,3,7,8-tetrachlorodibenzo-p-dioxin (2,3,7,8-TCDD) and related compounds on the uptake of intracellular calcium ([Ca2 +]i) in primary cultures of rat hippocampal neuronal cells. [Ca2 +]i levels were detected and quantified by interactive laser cytometry with microscopic image analysis. Cells were noninvasively labeled with fluo-3/AM and all experiments were conducted on cultured rat hippocampal neurons 14 days in culture. Treatment of cell cultures with 2,3,7,8-TCDD (10–100 nM) resulted in a rapid concentration-dependent increase in [Ca2 +]i associated with a decrease in mitochondrial membrane potential and activation of α-protein kinase C (α-PKC). In contrast, 1,2,3,4-TCDD, a weak Ah receptor agonist, had no effect on [Ca2 +]i at concentrations as high as 10 μM and similar results were also observed for 2,2′,5,5′-tetrachlorobiphenyl. Maximal [Ca2 +]i was observed within 30 s after addition of 2,3,7,8-TCDD and remained elevated (at higher concentrations) above resting levels for the duration of the experiment. This rapid increase in [Ca2 +]i was blocked by addition of EDTA (2 mM) to the external medium or by pretreatment of the cells with the calcium channel antagonist nifedipine (10 μM). However, pretreatment of the cells with 100 μM cycloheximide failed to block calcium uptake in neuronal cells. These data indicate that rat hippocampal neuronal cells are responsive to 2,3,7,8-TCDD; however, the mechanism is not associated with altered gene transcription and may involve cellular membrane targets.
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ABSTRACT: 2, 3, 7, 8-tetrachlorodibenzo-p-dioxin (TCDD) has been reported to cause alterations in cognitive and motor behavior during both development and adulthood. In this study, the neuronal nitric oxide synthase (nNOS) signaling pathway was investigated in differentiated pheochromocytoma (PC12) cells to better understand the mechanisms of TCDD-induced neurotoxicity. TCDD exposure induced a time- and dose-dependent increase in nNOS expression. High levels of nitric oxide (NO) production by nNOS activation induced mitochondrial cytochrome c (Cyt-c) release and down-regulation of Bcl-2. Additionally, TCDD increased the expression of active caspase-3 and significantly led to apoptosis in PC12 cells. However, these effects above could be effectively inhibited by the addition of 7-nitroindazole (7-NI), a highly selective nNOS inhibitor. Moreover, in the brain cortex of Sprague–Dawley (SD) rats, nNOS was also found to have certain relationship with TCDD-induced neuronal apoptosis. Together, our findings establish a role for nNOS as an enhancer of TCDD-induced apoptosis in PC12 cells.Environmental Toxicology and Pharmacology 07/2014; 38(1). DOI:10.1016/j.etap.2014.05.003 · 1.86 Impact Factor
Toxicological Sciences 01/2002; 69(1):191-201. DOI:10.1093/toxsci/69.1.191 · 4.48 Impact Factor
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ABSTRACT: The neurotoxic mechanism of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) has not been completely elucidated. In this study we investigated the possible role of cell cycle regulators and their dependence on arylhydrocarbon receptor (AhR) in the TCDD-mediated inhibition of cell proliferation using a human neuronal cell system. TCDD suppressed DNA synthesis of SK-N-SH human neuronal cells determined by [3H]thymidine incorporation which was significantly prevented either by pretreatment with α-naphthoflavone (α-NF), a partial AhR antagonist, or 8-methoxypsoralen (MOP), a binding inhibitor of activated AhR to dioxin response elements. Cell cycle analysis showed that TCDD induced a G1 cell cycle arrest, which was also significantly prevented by pretreatment with α-NF and MOP. TCDD did not alter the expression of cyclin D, cyclin E, p21 and p53. However, TCDD induced an enhanced expression of p27 and a hypophosphorylation of pRb, which was prevented by α-NF and MOP. Combined, these results suggest that the TCDD-induced inhibition of neuronal cell proliferation may be due to the AhR-dependent G1 arrest through an enhanced expression of p27 and a hypophosphorylation of pRB.Neuroscience Letters 06/2004; 363(1):69-72. DOI:10.1016/S0304-3940(04)00359-3 · 2.06 Impact Factor