The study was approved by the Ethics Committee of Affiliated Hospital of North Sichuan Medical College, Nanchong, China (Approved No. 2017/043). Written informed consents were obtained from all participants. All experiment procedures (consisted of invasive manipulation) and data collection were conducted with prior informed consents. This study adhered to the applicable CONSORT guidelines and was registered with the Chinese Clinical Trials Registry at http://www.chictr.org.cn (ChiCTR1800015819, principal investigator: Yanxia Guo, date of registration: April 23, 2018) .
The research was conducted between May 2018 and March 2019. Eighty five patients were American Society of Anesthesiologists (ASA) physical status Ⅰ-Ⅱ, aged between 30 and 65 years, and randomly assigned to five groups (S0, S1, S2, S3, S4) according to computer generated randomization. Patients in the five groups were anaesthetized by inhalation of sevoflurane and intravenous infusion of sufentanil with different plasma target concentrations (0.0, 0.1, 0.3, 0.5, 0.7 ng ml-1), respectively.
Exclusion criteria included patients with a history of cardiovascular, lung, kidney or brain disease; long-term drug and alcohol abuse; recent taking drugs known to affect the sympathetic adrenergic and cardiovascular systems; body mass index (BMI) ≧30 kg m-2. Withdrawal criteria included the patients with mean arterial pressure (MAP) <50 mmHg or heart rate (HR) < 50 bpm at any time during experimental observation; failing to the creation of carbon dioxide pneumoperitoneum for the first time or asking for adjustment of the pneumoperitoneal pressure above or below the preset value.
All patients were fasted at least for 8 h before surgery and without any preoperative medication. Before induction of anaesthesia, patient’s MAP, HR, electrocardiogram, pulse oxygen saturation were routinely monitored with a PM-9000 express monitor (Mindray Medical International Limited, Shenzhen, China). Simultaneously, a peripheral intravenous catheter was inserted for infusion of Ringer's solution with a rate of 10 ml kg-1 h-1. An arterial catheter was inserted into the left radial artery for
monitoring patient’s invasive arterial blood pressure and collecting blood samples. Anaesthesia was induced by inhalation of 8% sevoflurane with 100% oxygen until patients lost their consciousness, and 0.6 mg kg-1 of rocuronium was intravenously injected to facilitate laryngeal mask airway (Tuoren medical equipment group co. LTD, Henan, China ) insertion. Then mechanical ventilation was begun using 100% oxygen with a tidal volume of 6 to 8 ml kg−1 in each group. A normal end tidal carbon dioxide (CO2) tension (35 to 45 mmHg) was obtained by adjusting the respiratory frequency at 12 to 16 breaths min-1. The end-tidal sevoflurane concentration and CO2 partial pressure were monitored continuously using the above mentioned monitor.
Depth of anaesthesia was monitored by the bispectral index (BIS) (Canwell Medical International Limited, Zejiang, China) installed before induction.
Measurement of MACBAR
After laryngeal mask airway insertion, sufentanil was administered by target-controlled infusion with Bovil pharmacokinetic model using a micro pump (TCI-I, ver 4.0, Guangxi VERYARK Technology Co., Ltd), and the plasma target concentration of sufentanil was 0.0, 0.1, 0.3, 0.5, 0.7 ng ml-1 in groups S0, S1, S2, S3, S4, respectively. Simultaneously, the inhaled sevoflurane concentration was adjusted to obtain a stable preset end-tidal value according to our pilot study. In order to avoid a potential risk of intraoperative awareness, a higher initial end-tidal sevoflurane concentration was tested in the pilot study. Eventually, the first patient in S0, S1, S2, S3 and S4 group receiving a start end-tidal sevoflurane preset concentration of 5.0%, 4.6%, 3.0%, 2.3% and 2.0% which was measured to be close to the MACBAR, respectively. An up-and-down sequential-allocation method was applied to determine the MACBAR of sevoflurane in each group as described in our previous studies[11,12].
The CO2 pneumoperitoneum was created when the preset end-tidal sevoflurane concentration had been maintained stable at least 15 min. The creation of pneumoperitoneum was initiated using a Veress needle with CO2 to 13 mmHg at umbilicus and the insufflation flow rate was set at 3L/min. Then using a 10-mm trocar replaced the Veress needle. Another 10-mm trocar and 5-mm trocar were installed through subxiphoid port and a port in the right subcostal area of the midclavicular line, respectively. HR and MAP were determined before induction, 3 and 1 min before O2 pneumoperitoneum, 1 and 3 min after three trocars were installed. A positive or negative sympathetic adrenergic response to haemodynamic parameters (HR or MAP) was observed during the creation of CO2 pneumoperitoneum. The mean value of the MAP or HR measured 3 and 1 min before pneumoperitoneum stimulation was defined as its pre pneumoperitoneum value, and the mean value of HR or MAP measured 1 and 3 min after three trocars were installed was defined as the post pneumoperitoneum value. If the response was positive (an increase of patient’s HR or MAP over 20% of its pre pneumoperitoneum value), the subsequent tested patient’s end-tidal sevoflurane concentration would be increased by 0.2%, in contrast, If the response was negative (an increase of HR and MAP less than 20% of its pre pneumoperitoneum value), the subsequent tested patient’s end-tidal sevoflurane concentration would be decreased by 0.2%. Patients with bradycardia (HR<50bpm) or hypotension (MAP <50mmHg) at any time during experimental observation administered vascular active drugs such as atropine, ephedrine, would be withdrawn from the study, and the same concentration of sevoflurane was repeated in the following case. The study was continued until six crossing points of a negative versus positive response in the pre-and the next patient had occurred. The investigator responsible for recording the response of the patients to CO2 pneumoperitoneum was blinded to the plasma target controlled sufentanil concentrations and end-tidal sevoflurane concentration used in all the 5 groups. The MACBAR of sevoflurane in each group was calculated as the mean value of the end-tidal sevoflurane concentrations corresponding to the six crossing points.
After the above test was completed, the target controlled infusion of sufentanil was stopped in each group. The patients in group S0 received an i.v. bolus of 0.3 μg kg-1 sufentanil. Furthermore, the inspired concentration of sevoflurane was adjusted to maintain the end-tidal concentrations at 1.4-1.7 MAC for maintaining the BIS value between 40 and 60. MAP was maintained between 60 and 85 mmHg intraoperatively. If the MAP increased by more than 20% compared with its preoperative value, a bolus of 10 μg sufentanil would be administered. After removed laryngeal mask airway, patients were transported to post- anaesthesia care unit (PACU). In the PACU all patients were asked about whether there was an intraoperative awareness or not.
Analysis of blood samples
Arterial blood samples were collected 3 min before and after CO2 pneumoperitoneum with sodium-heparin-containing tubes. Soon after, the plasma was separated and kept frozen at -70 °C in a refrigerator until analysis. The method used to measure the concentrations of E and NE in the current investigation had been described previously.
Statistical analysis was performed using SPSS 23.0 software. The MACBAR was estimated from the up-and-down sequences using the method of independent paired reversals, which enabled MACBAR with 95%CIs to be derived. The sequences were also subjected to isotonic regression analyses. To compare the MACBAR from different groups more precisely, the 83% CIs were estimated using the isotonic regression analysis. The delta HR, delta MAP, delta E, delta NE value were calculated as the differences between their average values measured 1 and 3 min after CO2 pneumoperitoneum and before CO2 pneumoperitoneum, respectively. The data are presented as mean (SDs or 95%CI). The preoperative data, including gender and ASA class were compared with X2 test. The preoperative data (age, BMI), the intraoperative data, the postoperative data, the MACBARS, the concentrations of E and NE, delta E, delta NE, MAP, delta MAP, HR, delta HR were compared among the 5 groups using one-way analysis of variance (ANOVA). P value <0.05 was considered as statistical significance.