Long-term effects of recurrent neonatal seizures on neurobehavioral function and related gene expression and its intervention by inhibitor of cathepsin B.

Cathepsins are families of proteases that have been reported to play the key roles in neuroexcitotoxicity. The present study was sought to determine the effect of CBI, a cathepsin B inhibitor, in the prevention of neurobehavioral deficits after inhalant flurothyl-induced recurrent neonatal seizures in rats. We examined the expression pattern of autophagy-related genes at acute phase after the last seizures using western blot method, and evaluated behavioral deficits during postnatal day 28 (P28) to P35. The results showed improved neurological scores and learning ability in CBI-treated rats compared with the nontreated control. Flurothyl-induced increases in the ratio of LC3-II/LC3-I, Beclin-1 and Cathepsin-B were blocked by pre-treatment with CBI at 1.5, 3, 6 and 24 h after the last seizures in hippocampus and cerebral cortex by western blot analysis. Meanwhile, CBI also reversed flurothyl-induced down-regulation of Bcl-2 protein levels. Furthermore, in the long-term time point of 35 days (P35), PRG-1 mRNA and protein level in hippocampus and cerebral cortex of recurrent seizure group were up-regulated when compared to the control rats; meanwhile, the up-regulated expression of PRG-1 were robustly inhibited by CBI. These date demonstrated, for the first time, that lysosomal enzymes participate in neonatal seizure-induced brain damage and that modulation of cathepsin B may offer a new strategy for the development of therapeutic interventions for treatment of developmental seizure-induced brain damage.

Autophagy inhibitor 3-methyladenine regulates the expression of LC3, Beclin-1 and ZnTs in rat cerebral cortex following recurrent neonatal seizures.

BACKGROUND: Autophagy is a homeostatic process for intracellular recycling of bulk proteins and aging organelles. Increased autophagy has now been reported in experimental models of traumatic brain injury, stroke and excitotoxicity, and in patients with Alzheimer's disease and critical illness. The role of autophagy in developmental epilepsy, however, is unknown. The present study was to investigate the effects of recurrent neonatal seizure, in the presence and absence of autophagy inhibitor 3-methyladenine (3-MA), on the acute phase gene expression of ZnTs, LC3 and Beclin-1 in rat cerebral cortex and the interaction among them. METHODS: Thirty-six Sprague-Dawley neonatal rats at postnatal day 6(P6) were randomly divided into three groups: a recurrent-seizures group (RS, n=12), a 3-MA treated-seizure group (3-MA group, each rat pretreated with 3-methyladenine before seizures, 100nmol/µl/day, i.p., n=12) and a control group (n=12). At 1.5 and 6 hours after the last seizures, the mRNA levels of ZnT1-ZnT3, microtubule-associated protein 1A/1B light chain 3 (LC3) and beclin-1 were detected using the real-time RT-PCR method. The LC3 protein level was examined by Western blotting. RESULTS: The levels of LC3, beclin-1 and ZnT-2 transcripts in the RS group elevated significantly at 1.5 and 6 hours after the last seizures compared with those in the control and 3-MA groups. At the interval of 1.5 hours, the mRNA level of ZnT-1 increased significantly after the last seizure compared with that in the control group. There was no significant difference in the transcript levels of ZnT-3 among the three groups. Linear correlation analysis showed that the expression of the five genes in the control group exhibited a significant inter-relationship. In the 3-MA group, however, the inter-relationship was only found between beclin-1 and ZnT-1. In the RS group, the inter-relationship was not observed. CONCLUSIONS: The autophagy/lysosomal pathway is immediately activated along with the elevated expression of ZnT1 and ZnT2 in the cerebral cortex after recurrent seizures. 3-MA is involved in the regulation of the autophagy/lysosomal pathway and ZnTs by down-regulating the expression of LC3 and beclin-1.

[Observation on non-invasive electrode pulse electric stimulation for treatment of Bell's palsy].

OBJECTIVE: To explore non-invasive therapy for treatment of Bell palsy. METHODS: Two hundred and seventy-six were randomly divided into two groups, a treatment group and a control group, 138 cases in each group. The treatment group were treated with non-invasive electrode pulse electric stimulation at Taiyang (EX-HN 5), Sibai (ST 2), Qianzheng (Extra), Dicang (ST 4), and the control group with routine medicine (prednisone, dibazol, vitamine B complex and Qianzheng Powder), once each day, 10 days constituting one course. After two courses, their therapeutic effects were compared. RESULTS: The cured rate and the effective rate were 83.3% and 99.3% in the treatment group, and 48.5% and 88.4% in the control group respectively with a significant difference between the two groups (P < 0.05). CONCLUSION: Non-invasive electrode pulse electric stimulation at facial points has obvious therapeutic effect on Bell palsy.