Lead (Pb) is a well-known neurotoxicant and a risk factor for neurologic disorders. The blood brain barrier (BBB) plays an important role in the maintenance of optimal brain function. BBB is a target of Pb, and studies have shown that Pb induced barrier loss and decreased the expression of tight junction proteins, but the detailed mechanisms are not fully understood. Matrix metalloproteinases (MMPs) are important components of extracellular matrix proteasome and can affect the remodeling and degradation of tight junction (TJ). The role of MMP-2/9 in Pb-induced damage of BBB is not known. In our study, we used an in vitro BBB model by co-culturing human umbilical vascular endothelial cells (ECV304 cells) with rat glioma cells (C6 cells), and detected the expression of related TJ proteins and MMP-2/9. Our results showed that Pb increased the permeability of the in vitro BBB model, and stimulating C6 cells with Pb could decrease the protein level of ZO-1 (zonula occludens-1) and occludin in ECV304 cells. Pb could increase the mRNA and protein level of MMP-2/9 in C6 cells, and inhibition of MMP-2/9 by SB-3CT could partially alleviate Pb-induced down-regulation of TJ proteins in ECV304 cells and Pb-induced barrier damage in the in vitro BBB model. Our research established potential therapeutic targets for modulating and preserving optimal BBB function.

κ-opioid receptor (κ-OR) plays a key role in preventing hypoxic pulmonary hypertension (HPH) development after activated by exogenous agonist U50,488H. Calcium sensing receptor (CaSR) activation induces HPH by promoting vasoconstriction and vascular remodeling. The activated κ-OR is reported to inhibit the expression of CaSR in pulmonary artery smooth muscle cells (PASMCs). Thus, in this study, we aimed to explore the effect of activated κ-OR on the role of CaSR in preventing HPH development. An HPH rat model was constructed using Sprague-Dawley rats. Changes in mean pulmonary arterial pressure (mPAP) and right ventricular pressure (RVP) mediated by κ-OR agonist U50,488H and CaSR inhibitor NPS2143 were observed. The effects of CaSR agonist spermine and inhibitor NPS2143 on pulmonary artery tension were tested. The expression and localization of κ-OR and CaSR were measured in isolated PASMCs. A cell-counting kit-8 assay was performed to evaluate the effect of spermine in PASMC proliferation. Expression of proliferating cell nuclear antigen (PCNA), Erk, and p-Erk was evaluated by western blot analysis. Results showed that κ-OR and CaSR were co-expressed and colocalized in PASMCs under normoxic and hypoxic conditions. Interactions between κ-OR and CaSR were also observed. Spermine improved vasoconstriction in the pulmonary artery in HPH rats, which was abolished by U50,488H. RVP and mPAP were significantly increased in HPH rats under CaSR stimulation, but were significantly reduced when the rats were pretreated with U50,488H and NPS2143 (P < 0.01). Spermine treatment significantly promoted PASMC proliferation, which was significantly inhibited by U50,488H, p38 inhibitor SB203580, JNK inhibitor SP600125, Erk inhibitor SCH772984, and MEK inhibitor U0126, especially Erk inhibitor (P < 0.01). Spermine significantly increased PCNA and P-Erk expression in hypoxic conditions, which was inhibited by U50,488H and NPS2143. κ-OR stimulation prevented HPH development via the CaSR/MAPK signaling pathway.

Background: Lead and manganese are common neurotoxins. However, individuals are subject to co-exposures in real life, and it is therefore important to study these metals in combination. Methods: Weaning Sprague-Dawley rats were given ad libitum access to drinking water solutions containing lead (100 ppm), manganese (2.5 mg/mL) or a mixture, and each treatment has its own minocycline (50 mg/Kg.d) supplement group. Results: The results showed a significant difference in spatial memory and the induction levels of hippocampal long-term potentiation (LTP) in all exposure groups when compared with controls. The combined exposure group exhibited the most pronounced effect when compared with each of the single metal exposure groups. Microglia displayed activation at day 3 after exposure alone or in combination, while astrocytes showed activation at day 5, accompanied by decreased expression levels of glutamate/aspartate transporter (GLAST), glutamate transporter-1 (GLT-1), and glutamine synthetase (GS). Furthermore, the levels of the glutamate in the synaptic cleft increased significantly. When microglial activation was inhibited by minocycline, the activation of astrocytes, and the expression of GLAST, GLT-1, and GS were both reversed. In addition, upon minocycline treatment, the hippocampal LTP impairment and the cognitive injury were significantly alleviated in each of the exposure groups. Conclusions: These results suggest that combined exposure to lead and manganese can cause greater effects on cognition and synaptic plasticity when compared to single metal exposure groups. And the reason may involve in microglia abnormal activation leading to an excessive regulation of astrocytes, resulting in glutamate reuptake dysfunction in astrocytes and lead to perturbed cognition and synaptic plasticity.