Cardiac and sympathetic baroreflexes remain functional during local anesthesia with vasoconstrictor-free mepivacaine in humans

Local anesthesia with vasoconstrictor-free mepivacaine is known to not evoke pressor responses. However, it is unknown whether baroreflex function and blood pressure (BP) fluctuations are preserved by using mepivacaine. We tested the hypothesis that mepivacaine reduces baroreflex sensitivity (BRS) without changing its operating point. Beat-by-beat BP, heart rate (HR), and muscle sympathetic nerve activity (MSNA) were measured upon injection of either saline (CNT) or 3% mepivacaine (MPV) in the apical regions of the premolars and around the mandibular foramen in 10 healthy young men [23±5 (SD) years]. Cardiac and sympathetic BRSs were assessed by bolus injections of sodium nitroprusside followed by phenylephrine HCl, and then determined from the slopes of regression lines between systolic BP and HR and between diastolic BP and MSNA, respectively. HR was significantly higher in MPV than CNT (P<0.05), while there were no significant differences in MSNA, the operating points, or BP fluctuations between MPV and CNT (all P>0.05). Moreover, neither cardiac nor sympathetic BRS in CNT were altered by MPV (−0.71±0.13 vs. −0.78±0.33 beats·min−1·mmHg−1, P=0.41; −0.98±0.35 vs. −0.92±0.16 units・beat・ mmHg, P=0.73). Cardiac and sympathetic baroreflex functions were preserved and BP fluctuation may be well maintained under local anesthesia using vasoconstrictor-free 3% mepivacaine.


Introduction
Due to the medical development and population aging, the prevalence of cardiovascular diseases has increased, and the possibility of using local anesthesia for surgical procedures in patients with decreased cardiovascular reserve has been increasing. This has raised the risk of fatal cardiovascular events associated with local anesthesia, especially during dental treatments (1,2). Thus, it is important to evaluate the effects of local anesthetics on cardiovascular control systems for the safety of patients.
Most anesthetic solutions contain vasoconstrictors (e.g., adrenaline) to enhance the efficacy and prolong anesthesia duration. Despite their beneficial properties, it has been reported that administration of more than 40 µg adrenaline affects cardiovascular parameters (3). For example, the plasma adrenaline concentration reached 2-fold greater than resting levels after administration of 1.8 ml of lidocaine with 1:100,000 adrenaline (4), which was equivalent to that during moderate-to-heavy exercise (5). Therefore, its sympathoexcitatory effects may evoke cardiovascular mortality in patients at risk. The risks of using such vasoconstrictors should be weighed against the benefit, especially in cardiovascular patients.
Because mepivacaine, an amide-type local anesthetic, can sufficiently block somatosensory afferents and its half-life is long enough for almost all minor surgeries, such as dental procedures, even without vasoconstrictors, it is recommended for patients with cardiovascular diseases (6,7). Indeed, intraoral administration of 3% mepivacaine did not alter the average blood pressure (BP) per unit time in humans (8). However, the ability to maintain a steady BP should also be considered, because BP fluctuation beyond acceptable levels can cause cardiovascular events.
The baroreflex is a feedback system that maintains BP around its operating point (OP) by regulating cardiac output and total peripheral resistance through heart rate (HR) and muscle sympathetic nerve activity (MSNA) (9). The pressor responses to head-up tilt were greater in elderly blacks with lower baroreflex sensitivity (BRS) than in elderly whites (10), suggesting that blunted BRS contributes to greater BP responses during physiological stress.
Higher plasma adrenaline concentration (11) and local anesthetics itself such as lidocaine (12) and bupivacaine (13) were reported to reduce BRS in animal studies. However, the effects of mepivacaine on baroreflex functions are unknown.
We hypothesized that using mepivacaine decreases BRS without changing the OP.
Accordingly, we assessed the cardiac and sympathetic BRSs and the OPs by inducing BP alterations using intravenous injections of sodium nitroprusside (SNP) and phenylephrine HCl (PE) and compared responses to local anesthesia with 3% mepivacaine versus saline of the same volume.

Subjects
Ten healthy young men who were non-smokers with no overt history of cardiovascular or any other chronic diseases participated in this study. None were currently taking any medications. All subjects provided written informed consent to a protocol performed according to the Declaration of Helsinki and approved by the Institutional Review Board of Matsumoto Dental University (0266). Their physical characteristics (means±SD) were 24±4 years old, 176±6 cm tall, and 66.2±4.9 kg body weight.

Muscle sympathetic nerve activity:
MSNA signals were obtained using microneurography (14,15). In brief, a recording electrode (UNA35F4T; FHC, ME, USA) was placed in the peroneal nerve at the popliteal fossa, and a reference electrode was placed subcutaneously 2-3 cm apart from the recording 5 electrode. The nerve signals were amplified (70,000 to 160,000-fold), band-pass filtered (700 to 2000 Hz), full-wave rectified, and integrated with a resistance-capacitance circuit (time constant: 0.1 sec) to obtain a mean voltage neurogram. Criteria for adequate MSNA recording included pulse synchrony and facilitation during phase II of the Valsalva maneuver (15).

Hemodynamics:
HR was determined from the electrocardiogram (ECG) and arm cuff BP was measured by electronic sphygmomanometry with a microphone placed over the brachial artery on the left upper arm to detect Korotkoff sounds. Beat-by-beat BP was derived by finger photoplethysmography (Finometer; FMS, Amsterdam, The Netherlands), and the precision of beat-by-beat BP was verified by sphygmomanometry every minute.

Protocol
The experiment was performed in the morning ≥2 h after a light meal and ≥12 h after the last caffeinated or alcoholic beverage in a quiet, environmentally controlled chamber at an ambient temperature of ~29°C with a relative humidity of ~30%. The subjects were placed in the supine position and an 18-gauge Teflon catheter (SR-FS1832; TERUMO, Tokyo) was inserted into the antecubital vein for bolus injections of vasoactive drugs. After a satisfactory nerve recording site was found, the subjects rested for more than 20 min before starting the measurements. They then blindly underwent two sequential trials, first under placebo injections with saline (CNT), then second under an alveolar nerve block with 3% mepivacaine (MPV), which were separated by a 20-min rest. Each trial was consisted of a 5min baseline and 3-min measurements of HR and MSNA in response to vasoactive drugsinduced BP alterations. Throughout the entire procedure, respiratory rate was maintained at 6 16 times/min and the waveforms were continuously monitored by a piezo respiratory belt transducer (MLT1132, AD Instruments, Dunedin, New Zealand).

Local anesthesia with mepivacaine:
In MPV, we infiltrated 1.8 ml of 3% mepivacaine without vasoconstrictors into an area close to the mandibular foramen on the medial aspect of the mandibular ramus. We then injected another 1.8 ml of the solution into the buccal sides of the root apexes of upper and lower premolars. In CNT, 1.8 ml of saline solution was injected into each area in the same way as in MPV. The anesthetic effect was confirmed by stroking the gingival surface with a cotton ball and pricking it with a pin. If the depth of anesthesia was found to be insufficient, an additional amount of anesthetic solution was injected, but not beyond 300 mg or 4.4 mg/kg mepivacaine in subjects weighing <69 kg according to clinical practice guidelines (16).

Baroreflex test with vasoactive drugs:
The modified Oxford method was applied to determine cardiac and sympathetic BRSs, as reported previously (17). After baseline measurement, SNP (100 µg) was injected through the intravenous catheter to decrease BP. HR and MSNA responses were measured for the following 1 min. PE (150 µg) was then injected to increase BP and the responses were measured for the following 2 min.

Data analysis
Data were sampled at 625 Hz with a data acquisition system (PowerLab; ADInsturments, Dunedin, New Zealand). Offline data analyses were performed using signal-processing software (LabView; National Instruments, Austin, TX, USA). Beat-by-beat HR was calculated from the R-R interval. Beat-by-beat systolic BP (SBP) and diastolic BP (DBP) 7 were obtained from the arterial pressure waveform. SBP and DBP fluctuations were expressed by variance and the coefficient of variance (CV). Sympathetic bursts were identified from the integrated neurogram by a computer program (18) and confirmed by an experienced microneurographer who was blinded to the interventions.
The integrated neurogram was normalized by assigning a value of 1000 to the largest amplitude of a sympathetic burst during the 5-min baseline (9). Burst area was measured as the area under the curve of each sympathetic burst of the normalized integrated neurogram.
As quantitative indices, the number of bursts per minute (burst frequency), the number of bursts per 100 heartbeats (burst incidence), mean area per burst (mean burst area), and the area per beat (total MSNA) were calculated as follows:  (14,21). The pooling procedure reduces the statistical impact of inherent beat-by-beat variability attributable to non-baroreflex influence (e.g., respiration) (17). Moreover, a statistical weighting was adopted to minimize the effect of minor variations in bin position on the slope relative to the number of cardiac cycles in the bins (9).

Statistical analysis
Data analysis was performed using SigmaPlot12 (Systat Software Inc, Chicago, IL

Cardiovascular responses to vasoactive drugs
Representative changes in HR and integrated MSNA in response to altered BP by bolus injections of SNP and PE for a subject were shown in  (Figure 4C).

Discussion
The major findings were: the OPs of the baroreflex remained unchanged in SBP and DBP during MPV; baroreflex control of MSNA at the OP was not affected, while HR was reset to a higher level; cardiac and sympathetic BRSs were maintained; and BP fluctuations did not differ from those during CNT. Thus, using vasoconstrictor-free mepivacaine has little effects on cardiovascular control.
The OP is determined by the cardiovascular center in the medulla oblongata based on information from central command (22,23) and peripheral organs (e.g., chemoreceptor) (22)(23)(24)(25)(26)(27). In our study, as the same procedures were blindly applied to CNT and MPV, central command must be the same or affected equally between the conditions. Since the OP 11 remained unchanged by MPV, MPV did not have any peripheral effects on the OP. This was consistent with the previous reports demonstrating that MPV without vasoconstrictors did not alter average cuff BP (7,8,28,29), which should converge to the OP. In contrast, MPV increased HR, due to BP reduction through vasodilation elicited by MPV (30), which may be immediately compensated by increased HR via the cardiac baroreflex with an unaltered OP.
Higher HR increases oxygen demand of the heart, while vasodilation lowers afterload, resulting in little change in total cardiac workload. Thus, increased HR evoked by MPV with maintenance of the OP may not increase the cardiovascular risks, which was different from the simultaneous increases in HR and SBP developed with lidocaine with adrenaline (31) which is the most frequently used anesthetics for the minor surgeries.
Acute BP deviations from the safe range is one of the factors of cardiovascular events.
A higher BP variability was related to increased mortality (127%), and ~40% were more likely to have a heart attack or a stroke (32). Therefore, the ability to maintain BP within the safe range is important, as well as the average BP. If BRS is low, BP fluctuations increase and BP can frequently surpass the upper limit of safe levels. The use of lidocaine caused the reduction of cardiac BRS (12). Bupivacaine also depressed cardiac BRS in the presence of atenolol, but not in the presence of methylatropine in rats (13), indicating that the inhibition of cardiac BRS by bupivacaine may involve the vagal pathway. Conversely, cardiac BRS did not differ between MPV and CNT in our study, suggesting that mucosal infiltration of MPV maintains vagal control of HR. Since baroreflex modulation of HR is known to be mainly associated with SBP (17), the unaltered cardiac BRS may maintain SBP control during MPV. High BP, large BP fluctuations, and sympathetic nerve activation are independent cardiovascular event risk factors (36). Even if BP was controlled with antihypertensives, sympathetic nerve activity was higher than the normal (37). Therefore, such cardiovascular Limitations. First, it is possible that no difference in BRS between CNT and MPV may be a type II error due to a low statistical power. As no previous reports have investigated the effects of mepivacaine on cardiovascular variables, the minimum sample size was determined based on hemodynamic outcomes in a previous study evaluating the effects of lidocaine with adrenaline (8). Because the mean differences in BRSs were infinitesimal compared with the 95% CI, we believe that BRSs remained unchanged (or negligible) in MPV. Second, all subjects were men. Because BRS was reported to change with the menstrual cycle in women (38), we only included men to avoid any effects of the menstrual cycle. Third, we did not randomize the order of CNT and MPV because of the long-lasting effects of anesthesia. Since, we did not find any differences in hemodynamics, including BRS, the adaptation, fatigue, or central command in the second bout of baroreflex assessment was unlikely to affect BRS.
In summary, vasoconstrictor-free 3% mepivacaine has little or no impact on the cardiac and sympathetic BRSs or the OPs. The intact neurogenic BP regulation during local anesthesia with mepivacaine may be preferred in high-cardiovascular risk patients.

Data Availability
The original recorded waveforms and datasets generated during the current study are available from the corresponding author on reasonable request.      Values are means ± S.D. CNT, saline; MPV, 3%mepivacaine; SBP, systolic blood pressure; DBP, diastolic blood pressure; HR, heart rate; MSNA, muscle sympathetic nerve activity; *, P < 0.05.