Brain-derived neurotrophic factor produced long-term synaptic enhancement in the anterior cingulate cortex of adult mice.

Previous studies have demonstrated that brain-derived neurotrophic factor (BDNF) is one of the diffusible messengers for enhancing synaptic transmission in the hippocampus. Less information is available about the possible roles of BDNF in the anterior cingulate cortex (ACC). In the present study, we used 64-electrode array field recording system to investigate the effect of BDNF on ACC excitatory transmission. We found that BDNF enhanced synaptic responses in a dose-dependent manner in the ACC in C57/BL6 mice. The enhancement was long-lasting, and persisted for at least 3 h. In addition to the enhancement, BDNF also recruited inactive synaptic responses in the ACC. Bath application of the tropomyosin receptor kinase B (TrkB) receptor antagonist K252a blocked BDNF-induced enhancement. L-type voltage-gated calcium channels (L-VGCC), metabotropic glutamate receptors (mGluRs), but not NMDA receptors were required for BDNF-produced enhancement. Moreover, calcium-stimulated adenylyl cyclase subtype 1 (AC1) but not AC8 was essential for the enhancement. A selective AC1 inhibitor NB001 completely blocked the enhancement. Furthermore, BDNF-produced enhancement occluded theta burst stimulation (TBS) induced long-term potentiation (LTP), suggesting that they may share similar signaling mechanisms. Finally, the expression of BDNF-induced enhancement depends on postsynaptic incorporation of calcium-permeable AMPA receptors (CP-AMPARs) and protein kinase Mζ (PKMζ). Our results demonstrate that cortical BDNF may contribute to synaptic potentiation in the ACC.

Structural study of the novel deuterated calix[4]pyrrole complex d12-meso-tetrakis(4-methoxyphenyl)-meso-tetramethylcalix[4]pyrrole-pyridine N-oxide-acetonitrile (1/1/1).

Calix[4]pyrroles act as powerful receptors for electron-rich neutral guests and anionic guests in organic solvents. For the electron-rich neutral guest pyridine N-oxide, calix[4]pyrrole, with a deep cavity, provides an appropriate environment. The ability of calix[4]pyrrole to host binding guest molecules is the result of hydrogen bonding, π-π, C-H...π and hydrophobic interactions of the cavity. The novel title complex, C52H40D12N4O4·C5H5NO·C2H3N, based on d12-meso-tetrakis(4-methoxyphenyl)-meso-tetramethylcalix[4]pyrrole, has been assembled using an excess of pyridine N-oxide and is the first deuterated complex of calix[4]pyrrole. A single-crystal X-ray study shows that the receptor adopts a cone conformation with the N-oxide fragment encapsulated deep within the cavity. 1H NMR spectroscopy was used to probe the molecular binding formation in CD3CN. The results are consistent with the single-crystal X-ray study in identifying that the pyridine N-oxide molecule occupies the cavity of the calix[4]pyrrole molecule. UV-vis spectroscopy revealed that the calix[4]pyrrole receptor molecules are able to form 1:1 inclusion complexes in CH3CN.

[Influence of chronic fluorosis on the expression of mitochondrial fission protein dynamin-related 1 in the cortical neurons of rats].

OBJECTIVE: To explore the changes of protein expression of mitochondrial fission gene dynamin-related 1(Drp 1) in the cortical neurons of rats with chronic fluorosis. METHODS: A total of 120 one-month-old SD rats (each weighing approximately 100-120 g at the beginning of the experiment) were randomly divided into three groups, and fed with the different doses of fluoride containing in drinking water (untreated control containing 0 mg/L fluoride, and low-fluoride & high-fluoride supplemented with 10 and 50 mg/L fluoride,respectively). After 3 or 6 months exposure, 20 rats from each group were killed. Then the protein expression of mitochondrial fission gene, Drp1, was detected by immunohistochemistry and western-blotting method. RESULTS: Dental fluorosis and urinary fluorosis were obviously found in the rats exposed to fluoride. At the experiment period of 3 months, the numbers of positive cells of Drp1 detected by immunohistochemistry changed. Compared with the control group (36.3 ± 5.8), the changes in low-fluoride group (34.7 ± 4.1) showed no significant difference (t = 1.5, P > 0.05),but the increase in high-fluoride group (45.0 ± 4.7) had statistical significance (t = 8.8, P < 0.05). The western-blotting method had consistent results. Compared with the control group (0.59 ± 0.03), a significant increase of the average topical density in low- fluoride (0.62 ± 0.03) and high-fluoride (0.71 ± 0.02) groups were found (t = 0.02,0.11, P < 0.05). At the experiment period of 6 months, the numbers of positive cells of Drp1 detected by immunohistochemistry significantly changed. Compared with the control group (33.2 ± 4.4), the number in low- fluoride and high-fluoride groups were separately (36.6 ± 3.8) and (39.4 ± 4.2),both increased significantly (t = 3.5,6.3, P < 0.05). Same results could be found in western-blotting method,compared with the control group (0.65 ± 0.06), the average topical density in low- fluoride (0.80 ± 0.09) and high-fluoride (0.76 ± 0.08) groups both increased significantly (t = 0.1,0.1, P < 0.05). CONCLUSIONS: Taking excessive amount of fluoride might result in the changes of expression of Drp1, and the neurons damage from the chronic fluorosis might be associated with the hyperfunction of mitochondrial fusion.

[Changed expression of mito-fusion 1 and mitochondrial fragmentation in the cortical neurons of rats with chronic fluorosis].

OBJECTIVE: To observe the mitochondrial fragmentation and the expression of mito-fusion 1 gene in the cortical neurons of rats with chronic fluorosis, and to reveal their roles in mitochondria damage to neurons due to chronic fluorosis. METHODS: SD rats were divided randomly into three groups of 20 each (a half females and a half males housed individually in stainless-steel cages), and fed with the different doses of fluoride containing in drinking water (untreated control containing 0 mg/L fluoride, and low-fluoride and high supplemented with 10 and 50 mg/L fluoride, respectively). After 3 or 6 months exposure, the mitochondrial morphology of the neurons in rat brains were observed by transmission electron microscopy (TEM), then the expression of mitochondrial fusion gene, Mfn1, were detected by immunohistochemistry and western-blotting, respectively. RESULTS: Dental fluorosis was obvious in the rats exposed to excessive fluoride in their drinking water, that is, (16 rats out of 20) numbers of I° detal fluorosis in the low-fluoride group, and (11 rats out of 20) numbers of I° and (9 rats out of 20) numbers of II° detal fluorosis in the high-fluoride group were observed after 3 months exposure. Moreover, (14 rats out of 20) numbers of I° and (6 rats out of 20) numbers of II° detal fluorosis in the low-fluoride group and (6 rats out of 20) numbers of Io, (13 rats out of 20) numbers of II°, and (1 rats out of 20) numbers of III° detal fluorosis in the high-fluoride group were observed after 6 months exposure. And both of untreated controls without detal fluorosis were also observed. The urinary level of fluoride in the low-fluoride group (3.30 ± 1.18) mg/L and in the high-fluoride group (5.10 ± 0.35) were observed after 3 months exposure (F = 3.18, P < 0.05). Moreover, the urinary level of fluoride in the low-fluoride group (4.16 ± 1.39) mg/L and in the high-fluoride group (5.70 ± 1.70) mg/L were also observed after 6 months exposure (F = 3.17, P < 0.05). The normal mitochondrial morphology of neurons in rats without fluorosis was observed after 3 and 6 months, while the abnormal mitochondrial morphology of neurons with fluorosis was shown, presenting mitochondrial fragmentation with swollen cristae and even the fragmented, shortened or stacked punctuate membranes (section observation of three bullous mitochondrial-mitochondrial fission process) by TEM. As compared with controls (53.0 ± 4.54 and 1.21 ± 0.18) at the experiment period of 3 months, Mif1 protein analysis with immunocytochemical (the numbers of positive cells: 51.09 ± 6.25) and western-blotting (1.22 ± 0.26) were no significant difference for low fluoride group (t = 1.7, 1.1, P > 0.05); Mif1 protein analysis with immunocytochemical (the numbers of positive cells: 59.71 ± 5.64) and western-blotting (1.66 ± 0.20) were significantly increasing for high fluoride group (t = 2.1, 2.1, P < 0.05). As compared with controls (36.43 ± 4.04 and 1.00 ± 0.13) at the experiment period of 6 months, Mif1 protein analysis with immunocytochemical (the numbers of positive cells 20.05 ± 4.55 and 17.10 ± 3.86) and western-blotting (0.64 ± 0.08 and 0.39 ± 0.06) were significantly decreasing for the two fluoride group (t = 2.1, 2.2; 2.2, 2.2 respectively, all P value were < 0.05). CONCLUSIONS: Taking excessive amount of fluoride might result in the mitochondrial fragmentation for the changed expression of Mfn1, and the neurons damage from the chronic fluorosis might be associated with the dysfunction of mitochondrial fusion.

The influence of chronic fluorosis on mitochondrial dynamics morphology and distribution in cortical neurons of the rat brain.

The present study was designed to evaluate the effects of chronic fluorosis on the dynamics (including fusion and fission proteins), fragmentation, and distribution of mitochondria in the cortical neurons of the rat brain in an attempt to elucidate molecular mechanisms underlying the brain damage associated with excess accumulation of fluoride. Sixty Sprague-Dawley rats were divided randomly into three groups of 20 each, that is, the untreated control group (drinking water naturally containing <0.5 mg fluoride/l, NaF), the low-fluoride group (whose drinking water was supplemented with 10 mg fluoride/l) and the high-fluoride group (50 mg fluoride/l). After 6 months of exposure, the expression of mitofusin-1 (Mfn1), fission-1 (Fis1), and dynamin-related protein-1 (Drp1) at both the protein and mRNA levels were detected by Western blotting, immunohistochemistry, and real-time PCR, respectively. Moreover, mitochondrial morphology and distribution in neurons were observed by transmission electron or fluorescence microscopy. In the cortices of the brains of rats with chronic fluorosis, the level of Mfn1 protein was clearly reduced, whereas the levels of Fis1 and Drp1 were elevated. The alternations of expression of the mRNAs encoding all three of these proteins were almost the same as the corresponding changes at the protein levels. The mitochondria were fragmented and the redistributed away from the axons of the cortical neurons. These findings indicate that chronic fluorosis induces abnormal mitochondrial dynamics, which might in turn result in a high level of oxidative stress.

Protective action of endogenously generated H2S on hypoxia-induced respiratory suppression and its relation to antioxidation and down-regulation of c-fos mRNA in medullary slices of neonatal rats.

We previously reported that exogenous H(2)S played roles in protection of respiratory centers against hypoxic injury in medullary slices of neonatal rats. The protective action of endogenous H(2)S and its relation to antioxidation and down-regulation of c-fos mRNA were investigated in the present study. Perfusion of the slices with l-cysteine (Cys), substrate of cystathionine ß-synthase (CBS, H(2)S synthase), could increase frequency of rhythmic respiratory discharge of the hypoglossal rootlets and prevent respiratory suppression induced by hypoxia, whereas perfusion with hydroxylamine (NH(2)OH, inhibitor of CBS) could postpone recovery of respiration from hypoxic inhibition. NH(2)OH also significantly enhanced hypoxia-induced increase in malondialdehyde (MDA) content of the slices. The hypoxia-induced up-regulation of c-fos mRNA could be markedly antagonized by S-adenosyl-l-methionine (SAM, activator of CBS), but greatly increased by NH(2)OH. Neither NH(2)OH, Cys nor SAM had any effect on expression of bcl-2 mRNA in hypoxic medullary slices. These results indicate that endogenously generated H(2)S was involved in protection of the medullary respiratory centers against hypoxic injury partly via antioxidation and down-regulation of c-fos.

Protective effect of hydrogen sulfide on hypoxic respiratory suppression in medullary slice of neonatal rats.

Hydrogen sulfide (H(2)S) has been shown to play a protective role in injury of cells induced by hypoxia. Little is known however about its effect on medullary hypoxia-induced rhythmic respiratory suppression. In the present study, a decrease in frequency of rhythmic discharge of hypoglossal rootlets was observed in medullary slices of neonatal rats perfused with 95% N(2)-5% CO(2) to produce hypoxia. Perfusion with NaHS (H(2)S donor) prevented the inhibitory effect of hypoxia on the burst activity of the rootlets, whereas such action of NaHS was suppressed by pretreatment with glibenclamide, a blocker of K(ATP) channels. In addition, the increase in malondialdehyde content and the up-regulation of c-fos mRNA expression of the slices induced by hypoxia was significantly reduced by NaHS. These results indicate that exogenous H(2)S may protect the medullary respiratory center against hypoxic injury via activation of K(ATP) channels, reduction of lipid peroxidation and down-regulation of c-fos.