Caroline R. Stabernack

Gabaa Receptor Blockade Antagonizes the Immobilizing Action of Propofol but Not Ketamine or Isoflurane in a Dose-related Manner

The enhancing action of propofol on &ggr;-amino-n-butyric acid subtype A (GABAA) receptors purportedly underlies its anesthetic effects. However, a recent study found that a GABAA antagonist did not alter the capacity of propofol to depress the righting reflex. We examined whether the noncompetitive GABAA antagonist picrotoxin and the competitive GABAA antagonist gabazine affected a different anesthetic response, immobility in response to a noxious stimulus (a tail clamp in rats), produced by propofol. This effect was compared with that seen with ketamine and isoflurane. Picrotoxin increased the 50% effective dose (ED50) for propofol by approximately 379%; gabazine increased it by 362%, and both antagonists acted in a dose-related manner with no apparent ceiling effect (i.e., no limit). Picrotoxin maximally increased the ED50 for ketamine by approximately 40%–50%, whereas gabazine increased it by 50%–60%. The isoflurane minimum alveolar anesthetic concentration increased by approximately 60% with the picrotoxin and 70% with the gabazine infusion. The ED50 for propofol was also antagonized by strychnine, a non-GABAergic glycine receptor antagonist and convulsant, to determine whether excitation of the central nervous system by a non-GABAergic mechanism could account for the increases in propofol ED50 observed. Because strychnine only increased the immobilizing ED50 of propofol by approximately 50%, GABAA receptor antagonism accounted for the results seen with picrotoxin and gabazine. We conclude that GABAA antagonism can influence the ED50 for immobility of propofol and the non-GABAergic anesthetic ketamine, although to a different degree, reflecting physiologic antagonism for ketamine (i.e., an indirect effect via a modulatory effect on the neural circuitry underlying immobility) versus physiologic and pharmacologic antagonism for propofol (i.e., a direct effect by antagonism of propofol’s mechanism of action). This study also suggests that the immobilizing action of isoflurane probably does not involve the GABAA receptor because antagonism of GABAA receptors for animals anesthetized with isoflurane produces results quantitatively and qualitatively similar to ketamine and markedly different from propofol.

Glycine receptors mediate part of the immobility produced by inhaled anesthetics.

UNLABELLED Many inhaled anesthetics potentiate the effect of glycine on inhibitory strychnine-sensitive glycine receptors in vitro, supporting the view that this receptor could mediate the immobility produced by inhaled anesthetics during noxious stimulation (i.e., would underlie minimum alveolar anesthetic concentration [MAC]). There are quantitative differences between anesthetics in their capacity to potentiate glycines effect in receptor expression systems: halothane (most potentiation), isoflurane (intermediate), and cyclopropane (minimal). If glycine receptors mediate MAC, then their blockade in the spinal cord should increase the MAC of halothane more than that of isoflurane and isoflurane MAC more than cyclopropane MAC; the increases in MAC should be proportional to the receptor potentiation produced in vitro. Rats with chronically implanted intrathecal catheters were anesthetized with halothane, isoflurane, or cyclopropane. During intrathecal infusion of artificial cerebrospinal fluid, MAC was determined. Then MAC was re-determined during an infusion of 3, 12, 24, or 48 (isoflurane only) micro g/min of strychnine (strychnine blocks glycine receptors) in artificial cerebrospinal fluid. Strychnine infusion increased MAC in proportion to the enhancement of glycine receptors found in vitro. The maximum effect was with an infusion of 12 micro g/min. For the combined results at 12 and 24 micro g/min of strychnine, the increase in MAC correlated with the extent of in vitro potentiation (r(2) = 0.82). These results support the hypothesis that glycine receptors mediate part of the immobilization produced by inhaled anesthetics. IMPLICATIONS In vitro, halothane potentiates glycines effect on strychnine-sensitive glycine receptors more than isoflurane and isoflurane more than cyclopropane. The present in vivo work indicates that antagonism of the glycine receptor with strychnine increases minimum alveolar anesthetic concentration for halothane more than isoflurane and isoflurane more than cyclopropane. Such results support the notion that glycine receptors may mediate part of the immobility produced by inhaled anesthetics.

Gamma-aminobutyric acidA receptors do not mediate the immobility produced by isoflurane

Many inhaled anesthetics enhance the effect of the inhibitory neurotransmitter gamma aminobutyric acid (GABA), supporting the view that the GABAA receptor could mediate the capacity of inhaled anesthetics to produce immobility in the face of noxious stimulation (i.e., MAC, the minimum alveolar concentration required to suppress movement in response to a noxious stimulus in 50% of subjects). However, only limited in vivo data support the relevance of the GABAA receptor to MAC. In the present study we used two findings to test for the relevance of this receptor to immobilization for isoflurane: 1) differences among anesthetics in their capacity to enhance the response of receptor expression systems to GABA: isoflurane (considerable enhancement), xenon (minimal enhancement), and cyclopropane (minimal enhancement); and 2) studies showing that the spinal cord mediates MAC for isoflurane. If GABAA receptors mediate isoflurane MAC, then their blockade in the spinal cord should increase isoflurane MAC more than cyclopropane or xenon MAC and the MAC increase should be proportional to the in vitro enhancement of the GABAA receptor. To test this thesis, isoflurane, cyclopropane, or xenon MAC was determined in rats during intrathecal infusion of artificial cerebrospinal fluid (aCSF) via chronically implanted catheters. Then MAC was redetermined during infusion of 1 μL/min aCSF containing either 0.6 or 2.4 mg/mL picrotoxin, which noncompetitively blocks GABAA receptors. There was no consistent increase in MAC consequent to increasing the picrotoxin dose from 0.6 to 2.4 μg/min, which suggests that maximal blockade of GABAA receptors in the spinal cord had been achieved. Picrotoxin infusion increased MAC approximately 40% with all anes-thetics.ThisindicatesthatGABAreleaseinthespinalcord influences anesthetic requirement. However, the increase did not consistently differ among anesthetics and did not correlate with in vitro enhancement of GABAA receptors by these anesthetics. This supports the view that GABAA receptors do not mediate immobilization for isoflurane.

Absorbents differ enormously in their capacity to produce compound A and carbon monoxide.

Concern persists regarding the production of carbon monoxide (CO) and Compound A from the action of carbon dioxide (CO2) absorbents on desflurane and sevoflurane, respectively. We tested the capacity of eight different absorbents with various base compositions to produce CO and Compound A. We delivered desflurane through desiccated absorbents, and sevoflurane through desiccated and moist absorbents, then measured the resulting concentrations of CO from the former and Compound A from the latter. We also tested the CO2 absorbing capacity of each absorbent by using a model anesthetic system. We found that the presence of potassium hydroxide (KOH) and sodium hydroxide (NaOH) increased the production of CO from calcium hydroxide (Ca[OH]2) but did not consistently affect production of Compound A. However, the effect of KOH versus NaOH was not consistent in its impact on CO production. Furthermore, the effect of KOH versus NaOH versus Ca(OH)2 was inconsistent in its impact on Compound A production. Two absorbents (Amsorb® [Armstrong Medica, Ltd, Coleraine, Northern Ireland], composed of Ca(OH)2 plus 0.7% polyvinylpyrrolidine, calcium chloride, and calcium sulfate; and lithium hydroxide) produced dramatically lower concentrations of both CO and Compound A. Both produced minimal to no CO and only small concentrations of Compound A. The presence of polyvinylpyrrolidine, calcium chloride, and calcium sulfate in Amsorb® appears to have suppressed the production of toxic products. All absorbents had an adequate CO2 absorbing capacity greatest with lithium hydroxide. Implications Production of the toxic substances, carbon monoxide and Compound A, from anesthetic degradation by carbon dioxide absorbents, might be minimized by the use of one of two specific absorbents, Amsorb® (Armstrong Medica, Ltd., Coleraine, Northern Ireland) (calcium hydroxide which also includes 0.7% polyvinylpyrrolidine, calcium chloride, and calcium sulfate) or lithium hydroxide.

Spinal N-methyl-d-aspartate receptors may contribute to the immobilizing action of isoflurane.

We examined whether N-methyl-d-aspartate (NMDA) receptors influence the immobilizing effect of isoflurane by a spinal or supraspinal action. We antagonized NMDA receptors by intrathecal (IT), intracerebroventricular (ICV), and IV administration of MK 801 (a noncompetitive NMDA antagonist) and measured the decrease in isoflurane minimum alveolar anesthetic concentration (MAC). We also measured MK 801 tissue concentrations in homogenates of upper and lower spinal cord, a slice of cerebral cortex, and the whole brain. IT infusion of MK 801 decreased isoflurane MAC more potently than ICV or IV infusions. The change in MAC correlated with the MK 801 concentration in the lower part of the spinal cord (P < 0.01) but not with concentrations in supraspinal tissue. The maximal effect of IT MK 801 reached a plateau without achieving anesthesia. IV doses 270-fold larger than the largest IT dose also did not produce anesthesia in the absence of isoflurane. These results suggest that the capacity of MK 801 to decrease the MAC of isoflurane results from an effect on the spinal cord but that spinal NMDA receptors provide only partial mediation of the immobility produced by isoflurane. Because neither IT nor IV MK 801 provide complete anesthesia, these findings also call into question the notion that NMDA blockade alone suffices to produce anesthesia as defined by immobility in the face of noxious stimulation.

Temporal Summation Governs Part of the Minimum Alveolar Concentration of Isoflurane Anesthesia

Background General anesthesia may delay the onset of movement in response to noxious stimulation. The authors hypothesized that the production of immobility could involve depression of time-related processes involved in the generation of movement. Methods The delays (latencies) between onset of tail clamp (n = 16) or 50-Hz continuous electrical stimulation (n = 8) and movement were measured in rats equilibrated at 0.1–0.2% increasing steps of isoflurane. In other rats (n = 8), the isoflurane concentrations just permitting and preventing movement (crossover concentrations) in response to trains of 0.5-ms 50-V square-wave pulses of interstimulus intervals of 10, 3, 1, 0.3, or 0.1 s during the step increases were measured. These measures were again made during administration of intravenous MK801, an N-methyl-d-aspartate receptor antagonist that can block temporal summation (n = 6). Temporal summation refers to the cumulative effect of repeated stimuli. Crossover concentrations to 10- and 0.1-s interstimulus interval pulses ranging in voltage from 0.25–50 V were also measured (n = 4). Results The increase in concentrations from 0.6 to nearly 1.0 minimum alveolar concentration progressively increased latency from less than 1 s to 58 s. Shortening the interstimulus interval (50 V) pulses from 10 to 0.1 s progressively increased crossover concentrations from 0.6 to 1.0 minimum alveolar concentration. In contrast, during MK801 administration shortening interstimulus intervals did not change crossover concentrations, producing a flat response to change in the interstimulus interval. Increasing the voltage of interstimulus interval pulses increased the crossover concentrations but did not change the response to change in interstimulus intervals for pulses greater than 1 V. Conclusions Increasing the duration or frequency (interstimulus interval) of stimulation increases the concentration of isoflurane required to suppress movement by a 0.4 minimum alveolar concentration MK801 blocks this effect, a finding consistent with temporal summation (which requires intact N-methyl-d-aspartate receptor activity) at concentrations of up to 1 minimum alveolar concentration isoflurane.

Both cerebral GABAA receptors and spinal GABAA receptors modulate the capacity of isoflurane to produce immobility

We previously demonstrated that intrathecal administration of the noncompetitive &ggr;-aminobutyric acid type A (GABAA) receptor antagonist picrotoxin increased isoflurane MAC (the minimum alveolar concentration of anesthetic producing immobility in 50% of animals) by a maximum (ceiling effect) of approximately 40%. We also found that IV administration of picrotoxin increased MAC by more than 60%, without evidence of a ceiling effect. The larger increase with IV administration suggested a role of cerebral GABAA receptors. Accordingly, in this study we examined the effect of intracerebroventricular administration of picrotoxin in rats, finding that picrotoxin infusion into the third ventricle increased isoflurane MAC by a maximum of ap-proximately 40%, without finding a ceiling effect. In addition, we concurrently infused picrotoxin into the intrathecal and intracerebroventricular spaces, producing an increase in MAC in excess of 70%, also with no evidence of a ceiling effect. The dose-response relationship for the intrathecal-intraventricular infusion paralleled that of the IV infusion but was shifted to the left by an order of magnitude. We conclude that both cerebral and spinal GABAA receptors modulate the capacity of inhaled anesthetics to produce immobility. Because other studies have shown that the spinal cord, and not the brain, mediates the capacity of inhaled anesthetics to produce immobility, these results call into question the relevance of GABAA receptors to the immobilizing action of isoflurane.

Thiopental produces immobility primarily by supraspinal actions in rats.

The spinal cord mediates most of the immobilizing action of inhaled anesthetics. In the present study we investigated whether spinal or supraspinal sites mediate the immobilizing action of thiopental in rats. Thiopental was administered IV, intrathecally (IT), intracerebroventricularly (ICV), or simultaneously IT and ICV. Only the IV infusion produced anesthesia, defined as immobility in response to application of a tail clamp (i.e., the equivalent of minimum alveolar concentration, MAC). Consequently, the MAC-sparing effect (for isoflurane) of thiopental was used to assess the immobilizing contribution of IT and ICV infusions of thiopental. Thiopental concentrations were determined in whole brain, spinal cord, and a slice of cerebral cortex distant from the infusion sites. These concentrations were correlated with the MAC-sparing effect of the thiopental infusions in a multiple regression model. To assess the rate at which thiopental penetrates the cord, rat spinal cords were equilibrated in a bath of thiopental ex vivo and the concentration of thiopental in the cord was measured as a function of equilibration time. This was repeated in vivo with IT infusions of thiopental spanning the time of the behavioral studies. We found that IT or ICV infusion of thiopental 25 &mgr;g/min decreased isoflurane MAC <25%. The associated thiopental concentrations in the spinal cord after IT infusion, and in the whole brain after ICV infusion of 25 &mgr;g/min thiopental, exceeded by 500% and 680%, respectively, the concentrations found in the spinal cord and in the whole brain after IV infusion of thiopental in a dose that produced anesthesia in the absence of isoflurane. The percentage decrease in the MAC of isoflurane correlated primarily with the concentration of thiopental found in cerebral tissue not in contact with the cerebral ventricles. The spinal cord infusion produced an approximately 20% decrease in MAC. Ex vivo IT thiopental readily diffused into the spinal cord, with a time constant of approximately 1 h. We conclude that, unlike inhaled anesthetics, the immobilizing action of thiopental is largely supraspinal. Centers in the brain other than those near the third and fourth ventricles produce the greatest effect.

Blockade of 5-HT2A receptors may mediate or modulate part of the immobility produced by inhaled anesthetics.

Many inhaled anesthetics block the in vitro effect of the excitatory neurotransmitter serotonin on the 5-HT2A receptor, supporting the view that this receptor might mediate the capacity of inhaled anesthetics to produce immobility during noxious stimulation (i.e., would underlie MAC, the minimum alveolar concentration required to suppress movement in response to a noxious stimulus in 50% of subjects). In the present investigation in rats, we found that intrathecal administration of the 5HT-2A blocker, ketanserin, can decrease isoflurane MAC. This effect, presumably mediated by blockade of serotonin transmission in the spinal cord, reaches a maximum of 20%–25%. An additional decrease (to 60%) may be obtained by IV infusion of ketanserin, and presumably this decrease results from ketanserin’s actions on supraspinal centers. The IV doses of ketanserin that decreased MAC were approximately 100 &mgr;g · kg−1 · min−1 in rats, compared with usual clinical doses of 1.25 &mgr;g · kg−1 · min−1 in humans. These results indicate that 5HT2A receptors are in the neural circuitry influencing isoflurane MAC. These results, together with the blocking action of isoflurane on expressed 5HT2A receptors, strengthen the case for a role for 5HT2A receptors to isoflurane-induced immobility. However, because MAC for isoflurane is predominantly determined in the spinal cord, this result is consistent at most with a minor contribution of these receptors to the immobilizing action of isoflurane.

Sevoflurane degradation by carbon dioxide absorbents may produce more than one nephrotoxic compound in rats.

PurposeDegradation of sevoflurane by carbon dioxide absorbents produces compound A, a vinyl ether. In rats, compound A can produce renal corticomedullary necrosis. We tested whether other compounds produced by sevoflurane degradation also could produce corticomedullary necrosis.MethodsTwo groups of rats were exposed for four hours to sevoflurane 2.5% delivered through a container filled with fresh Sodasorb® and heated to 30∘C or to 50∘C, respectively. Compound A was added to produce an average concentration of 120 ppm in both groups. A third (control) group received 2.5% sevoflurane that did not pass through absorbent, and no compound A was added.ResultsAs determined by gas chromatography, the higher temperature produced more volatile breakdown products, including compound A. Median necrosis of the corticomedullary junction in the 50∘C group [10% (quartiles 1.096-7.8%); n = 20] exceeded that in the 30∘C group [5% (6.5%-15%); n = 18;P < 0.02], and both exceeded the median necrosis in the control group [0% (0.096-0.2%);n = 10;P < 0.02], The respective mean ± SD values for these three studies were: 12.8 ± 16.7%, 5.3 ± 4.4%, and 0.3 ± 0.5%.ConclusionDegradation products of sevoflurane other than compound A can cause or augment the renal injury in rats produced by compound A.RésuméObjectifLa dégradation du sévoflurane par les absorbants de gaz carbonique produit un éther vinylique, le composé A. Chez les rats, ce composé provoque une nécrose corticomédullaire rénale. Nous avons vérifié si d’autres composés issus de la dégradation du sévoflurane peuvent aussi provoquer cette nécrose.MéthodeDeux groupes de rats ont été exposés pendant quatre heures à du sévoflurane à 2,5 % administré après avoir traversé un récipient rempli de Sodasorb® frais et chauffé respectivement à 30° C ou à 50°C. Du composé A a été ajouté pour produire une concentration moyenne de 120 ppm dans les deux groupes. Un troisième groupe (témoin) a reçu du sévoflurane à 2,5 %, qui ne traversait pas l’absorbant, et sans ajout de composé A.RésultatsLes résultats de la Chromatographie en phase gazeuse ont montré que sous la température la plus élevée, il y a eu plus de produits de dégradation volatils, y compris le composé A. Dans le groupe 50°C, la nécrose moyenne de la jonction corticomédullaire dépassait [10 % (quartiles 1,0 %-7,8 %); n = 20] celle du groupe 30°C [5 %(6,5%- 15%);n = 18; P < 0,02] et les deux étaient plus élevée que celle du groupe témoin [0 % (0,0%-0,2 %); n = 10; P < 0,02]. Les valeurs respectives de la moyenne ± l’écart type ont été de 12,8 ± 16,7%, 5,3 ± 4,4 % et de 0,3 ± 0,5 %.ConclusionLes produits de dégradation du sévoflurane, autres que le composé A, peuvent causer ou augmenter la lésion rénale produite par le composé A chez les rats.