TASK Channels Contribute To Inhibition Function Of Sevoflurane To Surgical Stress In Mice

Part one Deletion of TASK channel changed inhibition function of sevoflurane to stress response in miceObjective:To set up the surgical stress model,and discuss whether the inhibition function of sevoflurane on surgical stress weakened in TASK-KO mice.Methods:32 wild C57BL/6 adult mice and 32 TASK-KO(Sichuan University Huaxi Hospital)C57BL/6 adult mice were selected(male).All mice were 8-10 weeks and 18-25g.Wild and TASK-KO mice were randomly respectively divided into 4 groups by random number group method.The wild type was divided into 4 groups(n=8):control group(Cl);sevoflurane group(S1);operation group(01);operation plus sevoflurane group(H1).The gene knockout type was divided into 4 groups:control group(C2),sevoflurane group(S2),operation group(02),operation plus sevoflurane group(H2).Intraperitoneal injection of 1%100 mg/kg pentobarbitol sodium before operation;control group with no conduction;S1 and S2 received simple sevoflurane inhalation for 10min after anesthesia,establishment of modified laparotomy surgical stress model after anesthesia in 01 and 02(completed within 10min),establishment of improved cesarean exploration model and sevoflurane inhalation(2.5%,oxygen flow rate 2L/min)for 10min in H1 and H2.After all experiments were finished,life monitoring of mice has been done at the same time in all groups:inclueding blood pressure,heart rate(IITC noninvasive blood pressure meter)and blood glucose(faster blood glucose meter),respectively.Results:In both wild group and TASK-KO group,the blood pressure,heart rate and blood glucose of the mice significantly risen after surgery compared with the control group(P<0.05);after the exposure of sevoflurane,the blood pressure and heart rate of mice in both wild group and TASK-KO group were significantly lower than those of the control group(P<0.05),while the blood glucose obviously declined only in wild mice(P<0.05);with the exposure of sevoflurane during surgery,the blood pressure and heart rate of the mice significantly declined compared with the control group(P<0.05),but the blood glucose only significantly desended in TASK-KO mice(P<0.05).In TASK-KO group,the basic state of blood pressure,heart rate and blood glucose was not changed,and the blood pressure and heart rate of the mice in other groups were both higher than those of the wild mice(P<0.05).Conclusion:Sevoflurane inhibited the surgical stress response in wild mice,and the inhibition effect weakened in TASK-KO mice.Part two Inhibition function of sevoflurane to surgical stress in mice via TASK channels in adrenal glandObjective:To confirmed that the inhibition function of sevoflurane in surgical stress whether be related to the TASK channel in adrenal gland.Methods:32 wild C57BL/6 adult mice and 32 TASK-KO(Sichuan University Huaxi Hospital)C57BL/6 adult mice were selected(male).All mice were 8-10 weeks and 18-25g.Wild and TASK-KO mice were randomly respectively divided into 4 groups by random number group method.The wild type was divided into 4 groups(n=8):control group(Cl);sevoflurane group(S1);operation group(01);operation plus sevoflurane group(Hl).The gene knockout type was divided into 4 groups:control group(C2),sevoflurane group(S2),operation group(02),operation plus sevoflurane group(H2).Intraperitoneal injection of 1%100 mg/kg pentobarbitol sodium before operation;control group with no conduction;S1 and S2 received simple sevoflurane inhalation for 10min after anesthesia,establishment of modified laparotomy surgical stress model after anesthesia in O1 and O2(completed within lOmin),establishment of improved cesarean exploration model and sevoflurane inhalation(2.5%,oxygen flow rate 2L/min)for 10min in H1 and H2.After all experiments were finished,eyeball blood,hypothalamus and adrenal tissue were obtained at the same time in all groups.The DNA and protein expressions of TASK channels in hypothalamus and adrenal gland of wild type and TASK-KO mice were detected by PCR and western blot.The concentration of corticotropin-releasing hormone(CRH),adrenocorticotropic hormone(ACTH)and serum corticosterone(CORT)were determined by enzyme-linked immunosorbent assay(ELISA).The concentration of adrenaline(E)and norepinephrine(NE)in serum were determined by high performance liquid chromatography(HPLC).Results:The contents of CRH,ACTH,plasma catecholamine and plasma corticosterone risen after surgery in the wild group and TASK-KO(P<0.05);after the exposure of sevoflurane,the contents of CRH,ACTH,plasma catecholamine and plasma corticosterone of the mice significantly declined(P<0.05).Compared TASK-KO mice of each group with the wild group,there was no significant difference in CRH and ACTH contents(P>0.05),while plasma catecholamine content and plasma corticosterone content were both higher than those of wild type mice(P<0.05).Conclusion:Inhibitory effect of sevoflurane on mice is mediated via TASK channels in adrenal gland.Part three Sevoflurane inhibited surgery stress via activating adrenal TASK channelsObjective:To study the effect of sevoflurane on TASK channel current and cell excitability in mice,and to further discuss its mechanism of the inhibition function of sevoflurane in surgical stress.Methods:8 wild C57BL/6 adult mice and 8 TASK-KO(Sichuan University Huaxi Hospital)C57BL/6 adult mice were selected(male).All mice were 8-10 weeks and 18-25g.To prepare the slice of adrenal gland,the full-cell patch-clamp technique was used to record the adrenal cell membrane voltage in resting state and the characteristic currents of TASK ion channels in adrenal cells at pH 5.9 and pH 7.3.The effect of sevoflurane on currents and excitability of adrenal slice cells was record and the relevant mechanism of this regulation in the surgical stress by sevoflurane was discussed.Results:In the resting state,the adrenocortical cell voltage was-74.3± 4.71 mV in wild mice,-54.8±3.65 mV in TASK-KO mice,the adrenal medullary cells voltage was-68.9±5.11 mV in wild mice,-58.7±3.85 mV in TASK-KO mice,the depolarization of TASK-KO mice showed more obviously than that of wild mice(P<0.05);in adrenocortical and adrenal medullary cell of wild mice,characteristic currents of TASK ion channels sensitive to pH were recorded,while pH-sensitive currents were absent in TASK-KO mice;in wild mice of this experiment,under the effect of sevoflurane,apparent change of adrenal and adrenal medullary cell current and significant increase of cell excitability were recorded,affecting the normal stress system,combined with the second part of the experimental results,which confirmed that sevoflurane led to the reduction of adrenal hormones;whereas in TASK-KO mice,the effect of sevoflurane did not cause significant fluctuations in the current of the cells,but led to the increase of adrenal hormones,indicating that the inhibition of sevoflurane on the stress system had been weakened in this moment.Conclusion:The patch clamp confirmed that sevoflurane inhibits surgical stress through the TASK channel in adrenal gland.