ABSTRACT
Objective:To observe and analyze the changes in activity of layer 2/3 cortical neurons in isoflurane-anesthetized mice by Real-time Ultra-large-Scale High-resolution (RUSH) imaging platform.Methods:Clean-grade healthy male Rasgrf2-Cre/Ai148d mice, aged 8-12 weeks, weighing 18-25 g, were studied.The mice recovered ten days after the skull replacement surgery and proceeded to the next experiment.Imaging data of calcium fluorescence signals from layer 2/3 cortical neurons were acquired by RUSH imaging platform after fixing the head of mice.The time of imaging data acquisition in the awake state, during anesthesia with 1.2% isoflurane, and after the end of anesthesia was 100, 600 and 600 s, respectively.Imaging data were analyzed using Image J and MATLAB softwares.Results:The overall trend of activity of layer 2/3 cortical neurons decreased first and then stabilized with the inhalation of 1.2% isoflurane.The cortical neural activity were gradually increased when isoflurane inhalation was stopped.The recovery rate of neural activity was different in different brain regions after isoflurane inhalation was stopped.The recovery of neural activity in the primary motor cortex was delayed obviously.During the maintenance of anesthesia, the activities of most layer 2/3 cortical neurons in the retrosplenial cortex were weakened, however, some of the neurons became more active.Conclusions:The neural activity in the 2/3 layer of cortex in isoflurane anesthetized mice is inconsistent in observation region, brain region and single cell, suggesting that different neural pathways are involved in the process of anesthesia induction and recovery from anesthesia.
ABSTRACT
Objective:To evaluate the role of Nod-like receptor family pyrin domain-containing 2 (NLRP2) in the dorsal root ganglion (DRG) in neuropathic pain (NP) in rats.Methods:Thirty-two male Sprague-Dawley rats in which intrathecal catheters were successfully implanted, aged 2-3 months, weighing 200-250 g, were divided into 4 groups ( n=8 each) using a random number table method: sham operation group (S group), NP group, NP plus NLRP2-siRNA group (NP+ siRNA group) and NP plus NLRP2-scrRNA group (NP+ scrRNA group). The right sciatic nerve was only exposed but not ligated in group S. NP was induced by chronic constriction injury (CCI) to the sciatic nerve in anesthetized rats in group NP, group NP+ siRNA and group NP+ scrRNA.NLRP2-siRNA and NLRP2-scrRNA were intrathecally injected at 3 days before CCI in group NP+ siRNA and group NP+ scrRNA, respectively.The mechanical paw withdrawal threshold (MWT) was measured at 1 day before CCI (T 0) and 1, 3, 7 and 10 days after CCI (T 1-4). The rats were sacrificed after the last measurement of pain threshold, and the L 4, 5 segments of the DRG on the operated side were removed for determination of the expression of NLRP2 and caspase-1 (by Western blot), the expression of NLRP2 mRNA (by real-time polymerase chain reaction) and interleukin-1beta (IL-1β) content (by enzyme-linked immunosorbent assay). Results:The MWT was significantly lower at T 2-4 than at T 0 in group NP, group NP+ siRNA and group NP+ scrRNA ( P<0.05). Compared with group S, the MWT was significantly decreased at T 2-4, the expression of NLRP2 protein and mRNA and caspase-1 was up-regulated, and the content of IL-1β was increased in group NP ( P<0.05). Compared with group NP, the MWT was significantly increased at T 2-4, the expression of NLRP2 protein and mRNA and caspase-1 was down-regulated, and the content IL-1β was decreased in group NP+ siRNA ( P<0.05), and no significant change was found in the parameters mentioned above in group NP+ scrRNA ( P>0.05). Conclusion:NLRP2 in DRG is involved in the development of NP, and the mechanism is related to NLPR2 inflammasomes-induced peripheral neuroinflammation in rats.
ABSTRACT
Combined with method of ketoconazole resistance screening, a 7alpha,15alpha-diOH-DHEA high-producing mutant Colletotrichum lini ST-1 was obtained by compound mutation of NTG and low energy N+ ion beam implantation. With the substrate concentration of 10 g/L DHEA, the molar yield of 7alpha,15alpha-diOH-DHEA reached 34.2%, increased by 46.2% than that of the original strain. Then we optimized the medium. First, Plackett-Burman design was used to evaluate the effects of medium components on molar yield of the product. Results show that glucose, yeast extract and MgSO4 x 7H2O were the important parameters for the biotransformation process. Subsequently, the path of steepest ascent was used to approach the optimal levels. To obtain the optimal levels, central composite design and response surface analysis were carried out. The optimal medium was as follows (g/L): glucose 26.34, yeast extract 12.15, corn flour 3.00, FeSO4 x 7H2O 0.015, MgSO4 x 7H2O 0.14, KH2PO4 0.90. Under the optimal conditions, the molar yield of 7alpha,15alpha-diOH-DHEA reached 49.3%, which was 44.2% higher than that of using the medium before optimization.