Lidocaine, MK-801, and MAC : Anesthesia & Analgesia

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Anesthetic Pharmacology: Research Report

Lidocaine, MK-801, and MAC

Zhang, Yi MD; Laster, Michael J. DVM*; Eger, Edmond I II MD*; Sharma, Manohar PhD*; Sonner, James M. MD*

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Anesthesia & Analgesia 104(5):p 1098-1102, May 2007. | DOI: 10.1213/01.ane.0000260318.60504.a9
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Lidocaine infusion decreases MAC. The largest decrease found varies as a function of study, dose of lidocaine, and animal. In humans, MAC decreases approximately 40% [determined as the effect in the presence of 70% nitrous oxide plus morphine premedication (1)]; in dogs the decrease is 37% (2) to 43% (3); in cats, 52% (4); in ponies by up to 70%, (5) and in rats, 50% (6). A maximum decrease of approximately 40%–50% appears to result, at least for some anesthetics and animals (1,6), a value similar to the 50%–60% maximum decrease produced by the blockade of N-methyl-d-aspartate (NMDA) receptors by dizocilpine (MK-801) (7). Is the mechanism underlying the decrease in MAC the same for MK-801 and lidocaine, i.e., blockade of transmission via NMDA receptors? Release of glutamate from nerve terminals may depend on the activity of sodium channels (8) and thus may determine the activity of NMDA receptors. Lidocaine, a local anesthetic that blocks sodium channels, might affect MAC by decreasing glutamate release, in effect doing what MK-801 does. Or it might directly block NMDA receptors (9).

The present study tested three ideas resulting from the above reasoning. First, as with MK-801, we predicted that a maximum decrease in MAC for various inhaled anesthetics will result from infusion of lidocaine, the decrease approximating that found with MK-801. Such a finding would indirectly support the hypothesis that the blockade of glutamate release mediates the effect of lidocaine. Second, if lidocaine and MK-801 produce the same net result (block of NMDA-mediated neurotransmission), then there should be other parallel findings. With conventional anesthetics, lidocaine should produce the previously described decrease of 50%–60% in MAC consequent to the infusion of lidocaine for conventional anesthetics but not for o-difluorobenzene which, at MAC, itself (alone) already blocks most NMDA receptors (7). That is, lidocaine infusion should have little effect on the MAC of o-difluorobenzene. Third, if lidocaine acts by limiting NMDA-mediated transmission, the addition of MK-801 should not decrease MAC more than the infusion of lidocaine alone, and should not (during lidocaine infusion) decrease the MAC of o-difluorobenzene as much as the MAC of conventional inhaled anesthetics.

METHODS

Materials

Isoflurane was obtained from Baxter Healthcare Corp. (New Providence, NJ); cyclopropane from Specialty Gases of America (Toledo, OH), halothane from Halocarbon (River Edge, NJ), and o-difluorobenzene from SynQuest Labs (Alachua, FL). Lidocaine was obtained from AstraZeneca LP (Wilmington, DE). MK-801 was obtained from Sigma-Aldrich (St. Louis, MO).

Studies of MAC in Rats

With approval of the Committee on Animal Research of the University of CA, San Francisco, we studied 93 male specific-pathogen-free, Sprague–Dawley rats (Crl: CD(SD)BR) weighing 300–450 g obtained from Charles River Laboratories (Hollister, CA). Each animal was caged with up to as many as two additional rats, and all had continuous access to standard rat chow and tap water before study. At least 24 h before study, IV catheters (PE 10 tubing, Portex, Hythe, Kent, England CT21 6JL) were placed in the right internal jugular vein under isoflurane anesthesia, and the open end of the catheter was tunneled to the ear where it exited and could be accessed.

MAC was determined concurrently in three to four rats placed in individual clear plastic cylinders. A rectal temperature probe was inserted, and the temperature probe and the tail of the rat were separately drawn through holes in the rubber stopper used to seal one end of the cylinder. Ports through the rubber stoppers in each end of the cylinder allowed gas delivery at the head end of the cylinder and exit of gas at the tail. For halothane and isoflurane, inflow rates of approximately 4 L/min were used. For cyclopropane and o-difluorobenzene, flow rates <1 L/min were used. For these anesthetics, the gases were recirculated through a carbon dioxide absorber, and the studies were conducted in a hood. An anesthetic concentration estimated to be less than MAC was administered for 40 min, after which the tail was clamped and moved for up to 1 min (less if the rat moved). After certifying that movement had occurred, the concentration was increased by 20%–25%, and after a 25–30-min period of equilibration the tail clamp was again applied and movement or lack of movement determined. This process continued until all rats failed to move in response to application of the tail clamp. During this MAC determination, saline was infused via the previously established IV catheter. MAC was calculated as the average of the largest concentration that permitted movement and the smallest concentration that suppressed movement.

Having established MAC, we started an infusion of lidocaine at 0 (control), 25, 100, 200, 300, 400, or 800 μg/min (only one infusion per rat) and restored the anesthetic concentration to one well below the MAC for the group. The decrease in inhaled anesthetic concentration was larger for greater infusions of lidocaine. After continuing lidocaine (or saline) administration for at least 50 min, we repeated the determination of MAC. When a given rat failed to move in response to stimulation, we opened the abdomen, cannulated the aorta with a 20 gauge catheter, and drew approximately 10 mL of arterial blood into a heparinized syringe (the exact volume was noted). Immediately after this exsanguination, the brain was removed and weighed.

Another series of rats were anesthetized with either isoflurane or o-difluorobenzene and MAC determined as above. A second MAC determination then was made in the presence of the administration of lidocaine at one of the infusion rates noted above plus the concurrent administration of 32 μg · kg–1 · min–1 of MK-801. As a given rat failed to move in response to noxious stimulation, 10 mL of arterial blood and the brain of the rat were removed and the brain weighed.

Extraction and Analysis of Lidocaine

The blood and brain samples obtained immediately after each rat failed to move in response to noxious stimulation were analyzed for their lidocaine content. The exact volume of blood removed and the weight of the brain were recorded. The brain was homogenized and 0.1 mL of a solution containing 10 mg/mL fluoride and buffer 1 mL 0.5 M sodium carbonate (pH 10.8) was added. We added a volume of n-pentane equal to twice the volume of the blood or brain, vortexed the mixture for 60 s, centrifuged the mixture, and removed and saved the supernatant pentane phase. Extraction of the lidocaine with n-pentane was repeated and the pentane added to the first volume of pentane. The pooled pentane was evaporated under nitrogen and the dried samples stored at −80°C until analyzed.

A model 1100 high-performance liquid chromatograph (Agilent 1100 series, Agilent Technologies, Mountain View, CA), equipped with an automatic sampling system was used. Analyses were performed using a 3.5 μm; 150 mm × 2.1 mm Agilent Zorbax Eclipse XDB-C18 column (Agilent Technologies) at ambient room temperature (20°C–25°C).

We used acetonitrile, methanol, and 0.05 M dibasic sodium phosphate (25:20:55) with a pH of 8.1 as the mobile phase. The flow rate was 0.25 mL/min and the eluant was monitored at 242 nm. The frozen samples were reconstituted in 100 or 200 μL of eluant, and a 40 μL aliquot was injected. Quantitation was performed by reference to a calibration curve of 0–200 μg/mL lidocaine in both blank blood and homogenized brain. Areas under the peak were measured and were correlated with the known 0–200 μg/mL concentrations with r2 > 0.99.

Analysis of Inhaled Anesthetics

We used a Gow-Mac gas chromatograph (Gow-Mac Instrument, Bridgewater, NJ) equipped with a flame ionization detector to measure concentrations of carbon-containing compounds. The 4.6-m long, 0.22 cm (ID) column was packed with SF-96. The column temperature was 100°C–200°C. The detector was maintained at temperatures approximately 50°C warmer than the column. The carrier gas flow was nitrogen at a flow of 15–20 mL/min. The detector received 35–38 mL/min hydrogen and 240–320 mL/min air. Primary standards were prepared for each compound, and the linearity of the response of the chromatograph was determined. We also commonly used secondary (cylinder) standards referenced to primary standards for some anesthetics (e.g., isoflurane).

Statistical Analyses

MAC for each rat was determined twice; once as a control (A), and a second time during infusion of lidocaine (B). The decrease in MAC for each rat was taken as (AB)/A. For a given anesthetic, these values were correlated with the infusion rate, and the blood and brain lidocaine concentrations specifically determined for each rat, and with the specific infusion rate of lidocaine. We used an analysis of variance to determine whether the decrease differed as a function of the inhaled anesthetic.

RESULTS

MAC correlated inversely with the infusion rate of lidocaine (Fig. 1). Data for the 800 μg/min infusion probably underestimated its effect on MAC because several rats did not move at this infusion rate despite discontinuing inhaled anesthetic delivery. Occasionally, a rat convulsed, and data from such rats were not included. For o-difluorobenzene, 4 of 9 rats died at an infusion rate of 400 μg/min before MAC could be determined and the data for this infusion rate also probably underestimated the effect on MAC. MAC correlated with either blood (Fig. 2) or cerebral (Fig. 3) concentrations of lidocaine. Concurrent administration of MK-801 and lidocaine produced effects on the MAC of isoflurane versus o-difluorobenzene that did not differ statistically (Fig. 4).

F1-17
Figure 1.:
Infusion of lidocaine decreased MAC in a dose-related manner for cyclopropane, halothane, isoflurane, and o-difluorobenzene. In this and the succeeding figures, the first three (conventional) anesthetics are represented by filled symbols; o-difluorobenzene is represented by open squares. Each point represents the data for 3–4 rats and provides the mean and sd for that point in this and all succeeding figures. The curves for the different anesthetics differ in position (ANOVA) but no one anesthetic differed significantly from the others (post hoc analysis). For clarity, the values are offset slightly from the true infusion rate values. At the greatest infusion rate, some rats required no inhaled anesthetic.
F2-17
Figure 2.:
Blood plasma lidocaine concentrations increased with increasing infusion rates and the decrease in MAC correlated with the increased plasma concentrations. Again, no one anesthetic seemed unequally affected. In particular, o-difluorobenzene was not differently affected.
F3-17
Figure 3.:
Whole brain lidocaine concentrations increased with increasing infusion rates and the decrease in MAC correlated with the increased brain concentrations. Again, no one anesthetic seemed unequally affected. In particular, o-difluorobenzene was not differently affected.
F4-17
Figure 4.:
The addition of 32 μg · kg–1 · min–1 MK-801 to the infusion of lidocaine appeared to equally decrease the MAC of isoflurane and o-difluorobenzene. The combination of MK-801 with infusions of 200 or 400 μg/min of lidocaine often alone was sufficient to produce immobility.

A two-way ANOVA examining the effect of infusion rate and choice of anesthetic on MAC revealed significant effects for both (choice of anesthetic produces an F of 3.9 and P = 0.016; inflow rate produces an F of 30.9 and P < 0.001). However, a post hoc analysis did not indicate which anesthetic differed significantly from other anesthetics. A multiple regression analysis with anesthetic choice and blood or brain concentration as variables did not reveal a significant difference among anesthetics. It appeared that lower infusion rates of lidocaine affected the MAC of o-difluorobenzene less than isoflurane (Fig. 4).

DISCUSSIONS

Our results failed to confirm any of the three hypotheses that prompted our study. First, we did not find a floor (maximum effect) on MAC resulting from increasing doses of lidocaine. Instead, we found a progressive decrease in the MAC for cyclopropane, halothane, isoflurane, and o-difluorobenzene (Figs. 1–4), one that went well beyond 40%–60%. In several rats given the greatest infusion rate of lidocaine, the decrease in MAC approached 100%. Thus, the effect of lidocaine on MAC would not appear to be solely due to blockade of release of NMDA.

Second, a given infusion of lidocaine did not decrease the MAC values for o-difluorobenzene less than the MAC values for the remaining anesthetics. This further indicates that lidocaine does not act primarily through a capacity to suppress the release of glutamate at nerve terminals.

Third, administration of MK-801 shifted the lidocaine dose–response relationship equally for o-difluorobenzene and isoflurane. Again, this would not be predicted if lidocaine acted in large part through inhibition of release of glutamate.

To recapitulate, the effect of lidocaine on MAC is essentially the same for halothane and isoflurane versus cyclopropane and o-difluorobenzene. Cyclopropane and o-difluorobenzene strongly block NMDAreceptors at MAC, whereas halothane and isoflurane weakly block these receptors (10). Previously this difference in blocking was shown to be associated with a different effect of MK-801 on the MAC of isoflurane versus o-difluorobenzene (but not cyclopropane) (7). The present study shows no such distinguishing effect on the action of lidocaine, further supporting a view that lidocaine must do more than simply block transmission through NMDA receptors.

The results of the present study do not reveal the basis for this additional or alternative effect of lidocaine. The systemic infusion of lidocaine, and, by implication, sodium channel blockade [but potassium channel effects may contribute, too (11)], may produce immediate and long-term decreases in nociception from surgery (12–16). Indeed, lidocaine might have diverse, yet to be defined effects on MAC. For example, lidocaine can block γ-aminobutyric acida, (17) glycine (18), and acetylcholine (19) receptors, and such combined effects might influence MAC.

Finally, the present data support the notion that administration of lidocaine might usefully supplement the clinical delivery of anesthesia. The decrease in MAC can be substantial. The postoperative analgesia associated with lidocaine's intraoperative administration compliments this action (12–16). Further advantages to the clinical intraoperative use of lidocaine include the protection of vital tissues, such as the brain, against periods of hypoxia (20), perhaps by activation of mitochondrial potassium ATP channels (21).

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