Transdermal Nitroglycerine Enhances Spinal Neostigmine Postoperative Analgesia following Gynecological Surgery

The Ethical Committee of the Teaching Hospital of the University of São Paulo, Ribeirão Preto, approved the study protocol. After giving informed consent, 48 patients, American Society of Anesthesiologists physical status I and II, scheduled to undergo vaginoplasty were randomized with use of a computer to one of four groups and prospectively studied using a placebo-controlled double-blind design to evaluate analgesia and adverse effects. The concept of visual analog scale (VAS), which consisted of a 10-cm line, with 0 equaling “no nausea” or “no pain at all” and 10 equaling “worst possible nausea” or “the worst possible pain” was introduced before surgery.

Patients were premedicated with use of 0.05–0.1 mg/kg intravenous midazolam in the holding room. Hydration consisted of 10 ml/kg Ringer’s lactate solution preoperatively and 10 ml · kg⁻¹· h⁻¹after spinal anesthesia. Spinal anesthesia was performed in the operating room at the L3–L4 interspace, with the patient in the sitting position. A volume of 4 ml was injected over 30 s through a 25-gauge spinal needle. The intrathecal drugs included 15 mg hyperbaric bupivacaine, 0.5% (3 ml), plus the test drug (1 ml). Patients were placed in the supine position immediately after spinal injection. The transdermal patch was applied at the thorax (ventral, between T2–T4), in a nonanesthetized area, 20–30 min after the spinal puncture (after hemodynamic stabilization). The drugs (intrathecal and transdermal) were prepared by one anesthesiologist. A second anesthesiologist who was blind to the drugs selected performed the lumbar puncture and drug injection, stayed during the intraoperative period, and checked postoperative analgesia and adverse effects. The groups (control, neostigmine, nitroglycerin, and neostigmine–nitroglycerin) are described in table 1.

Intraoperative sensory loss assessment included the pin prick test 10 min after spinal injection. Blood pressure was monitored noninvasively every 5 min throughout surgery, and heart rate and oxyhemoglobin saturation (Spo²) were monitored continuously during surgery. A decrease in mean arterial pressure greater than 15% below the preanesthetic baseline value was treated by incremental doses of ephedrine, 4 mg intravenously. Decreases in heart rate below 50 beats/min were treated with incremental doses of atropine, 0.25 mg intravenously. Intraoperative nausea was scored by the patient using the 10-cm VAS N (nausea). The number of patients having nausea (of any degree) or vomiting at any point intraoperatively was noted. Nausea greater than 2 cm on a 10 cm scale at any time or vomiting during the study were treated initially with 10 mg intravenous metoclopramide, followed by 0.5 mg intravenous droperidol, if necessary. For patients who had more than one episode of nausea, the VAS scores were averaged.

Postoperative assessment included pain scores, adverse effects and the duration of motor block, measured from anesthetic injection until the time to reach a Bromage score of 2. 10Patients were allowed to receive rescue analgesics, and there was always someone from the staff available to administer the analgesic at the time requested. Intramuscular diclofenac (75 mg) was available. Pain was assessed at the time of administration of first rescue analgesic and 24 h after the spinal puncture by the anesthesiologist who was blind to the treatment. Nausea and occurrence of vomiting were continuously assessed intraoperatively and 24 h after the spinal puncture by the same anesthesiologist, blind to the treatment. Duration of effective analgesia was measured as time from the intrathecal drug administration to the patient’s first request for analgesic administration, either in the recovery room or the infirmary, and was recorded in minutes. The VAS at the time of administration of first rescue analgesic medication was measured using the 10-cm VAS. The 24-h VAS pain score and VAS for nausea reflected the patient’s overall impression of the 24 h after spinal injection.

The number of subjects was based on preliminary experimental data. We hypothesized that intrathecal neostigmine would increase the time to administration of first rescue analgesic by 20% in the population studied and that the association of transdermal nitroglycerin patch would increase the time to administration of first rescue analgesic by 100% when compared with the control group. If we estimated a standard deviation for this prospective power analysis as 40% and an α value of 0.05, these assumptions would necessitate inclusion of five patients in each group to show a 100% increase in the time to administration of the first rescue analgesic. To further increase the power, we elected to observe 12 patients in each group.

The normality of the distributions was assessed using the Shapiro-Wilk test. Groups were compared with regard to demographic data (age, weight, height) and duration of surgery with use of one-way analysis of variance. Incidence of adverse events, gender, American Society of Anesthesiologists physical status, and adjuvant drug use were compared among groups with use of the chi-square test corrected for multiple comparisons. Probability was considered to be significant if less than 0.0125. Blood pressure, heart rate, level of anesthesia (pin prick), and VAS scores were compared among groups by two-way analysis of variance for repeated measures. 11Tukey analysis was applied to decrease the probability of type I error. The time to administration of first rescue analgesics was compared using the Kruskal-Wallis test, applied with a nonparametric multiple comparison procedure, and the Wilcoxon rank sum test. Time to administration of first rescue analgesics is expressed as the median (25–75th percentile). P < 0.05 was considered to be significant. Data are expressed as mean ± SD, unless otherwise stated.

The four groups showed no differences regarding American Society of Anesthesiologists physical status, age, weight, and height (P > 0.05;table 2). The sensory level to pin prick at 10 min, surgical and anesthetic time, and intraoperative ephedrine consumption were similar among groups (table 3).

The postoperative data are represented in table 4. The pain VAS score at the time of administration of first rescue analgesic medication was similar among the four groups (P > 0.05). The time to administration of first rescue analgesic medication (min) was longer in the neostigmine–nitroglycerin group compared with the other groups (P < 0.001). The analgesic consumption during the first 24 h postoperatively was less for the neostigmine–nitroglycerin compared with the control group (P < 0.0005) and also less for the neostigmine group compared with the control group (P < 0.02). The analgesic consumption of the other groups were similar.

There were no differences regarding the incidence of perioperative adverse effects (P > 0.05). Intraoperatively, none of the patients reported nausea or vomiting. Postoperatively, one patient from the control group reported back pain (VAS 5 cm) and another experienced one episode of vomiting. One patient from the nitroglycerine group reported a headache (VAS 3 cm); another reported back pain (VAS 4 cm); and one other experienced one episode of vomiting. One patient from the neostigmine group reported back pain (VAS 5 cm); another patient reported pain in the knee (VAS 4 cm); and another reported flatulence. Two patients from the neostigmine–nitroglycerin group vomited. The mean overall 24-h nausea VAS score was similar among groups (P > 0.05).

The results of this study showed that, although neither 5 μg spinal neostigmine nor transdermal nitroglycerine alone delayed the time to administration of first rescue analgesic, the association of 5 mg/day transdermal nitroglycerin patch and intrathecal low-dose neostigmine (5 μg) resulted in an average of 14 h of postoperative analgesia after vaginoplasty during bupivacaine spinal block, compared with 3.5 h in the control group. Nevertheless, the lack of a direct action on the time to administration of first rescue analgesic, patients who received 5 μg intrathecal neostigmine only as analgesic, had a lower analgesic consumption during the first 24 h of observation, compared with the control group. This lower consumption may reflect some analgesic effect of the dose used in the population studied.

Previously, we demonstrated that spinal neostigmine causes dose-dependent analgesia in patients undergoing vaginoplasty. 1Intrathecal neostigmine doses varying from 10 μg to 200 μg were limited by side-effects (namely, nausea and vomiting) when the doses varied between 25 and 200 μg. 2,3,12In addition, intrathecal neostigmine was more efficient for somatic rather than visceral pain, 13and the female population may have an advantage with neostigmine analgesia because of the sex difference in spinal cholinergic analgesic nicotinic mechanisms. 7All data directed us to further study female patients undergoing a purely somatic painful stimuli, i.e. , vaginoplasty surgery.

Intrathecal neostigmine antinociception is secondary to acetylcholine release and action in the spinal cord tissue. 4,5During surgical stimuli, a preexistent spinal cholinergic tonus is activated. 14The presence of acetylcholine in the cerebrospinal fluid has been shown in humans. 15Acetylcholine from this physiologic cholinergic mechanism and acetylcholine preserved from cholinesterase activity after intrathecal neostigmine will bind to muscarinic 7and nicotinic 8nerve terminals in the spinal cord. The remaining acetylcholine that reaches the cerebrospinal fluid is also preserved from cholinesterase activity located within the spinal meninges. 6This would increase acetylcholine cerebrospinal fluid concentration and improve acetylcholine bioavailability at cholinergic nerves within the spinal cord.

The transdermal nitroglycerin patch has been related to nitric oxide formation during degradation of organic nitrates. 16In accordance to animal 17and clinical research, 18nitric oxide generators did not result in analgesia. Nevertheless, a current study provides evidence that acetylcholine stimulate nitric oxide synthesis in the spinal cord, 9and this synthesis is necessary for the expression of analgesia secondary to the cholinomimetic agents, 19such as spinal neostigmine, as much as behavioral analgesia from intrathecal injection of muscarinic agonists in rats is inhibited by nitric oxide synthase blockers. 20 

In addition, the activation of descending pain pathways involves the participation of nitric oxide, which mechanisms of action are likely to include activation of second messengers such as cyclic guanosine monophosphate (cGMP). 21Wide-dynamic-range neurons in the superficial dorsal horn and high-threshold cells in the superficial or deep layers show reduced response after exposure to cyclic guanosine monophosphate. 22Therefore, analgesia would be a result of predominant analgesic action on superficial spinal layers.

Anatomic evidence also supports the connection between nitric oxide and acetylcholine. Nitric oxide synthase colocalizes in dorsal horn neurons that contain choline acetyl transferase. 23Nitric oxide synthase is localized to the superficial dorsal horn and the intermediolateral cell column regions of the spinal cord. 24Muscarinic receptors have been identified on spinal cord dorsal horn and intermediolateral cell columns. 25 

In conclusion, although intrathecal neostigmine alone (5 μg) or transdermal nitroglycerine alone (5 mg/day) (a nitric oxide generator) did not delay the time to administration of first rescue analgesic, the association of both provided 14 h of postoperative analgesia after vaginoplasty surgery, suggesting that nitric oxide and cholinergic receptors may enhance each other’s antinociceptive effects at the dose studied.