Mechanisms of Bronchoprotection by Anesthetic Induction Agents : Propofol versus Ketamine

Our study protocol was approved by The Johns Hopkins Animal Care and Use Committee. Anesthesia was induced in eight sheep (25-35 kg) with intramuscularly administered ketamine (30 mg/kg) and subsequently maintained with pentobarbital sodium (20 mg [middle dot] kg^-1[middle dot] h^-1). A tracheostomy was performed, the sheep were paralyzed with pancuronium bromide (2 mg intravenously, with supplementation during the experiment), and the lungs were mechanically ventilated with room air with supplemental oxygen at a rate of 15 breaths/min and a tidal volume of 12 ml/kg. Five centimeters of H²O positive end-expiratory pressure was applied. The left thorax was opened at the fifth intercostal space, and heparin (20,000 U) was administered. The esophageal and thoracic tracheal branches of the bronchoesophageal artery were ligated as previously described. [27]The bronchial branch was then cannulated with an 18-gauge angiocatheter and perfused with a constant flow (0.6 ml [middle dot] min^-1[middle dot] kg (^-1)) of autologous blood withdrawn from a femoral artery catheter by a variable-speed pump (Gilson, Villiers-Le-Bel, France). Systemic blood pressure, heart rate, and bronchial arterial pressure were measured continuously throughout the study.

Conducting airways resistance (R^aw) was measured by forced oscillation. [28]In this method, a gas volume of [almost equal to] 30 ml is oscillated for 1.5 s at a frequency of 9 Hz after each tidal breath. Airway pressure is measured at a side arm of the tracheal cannula, and a flow signal is obtained from a pneumotachograph positioned between the oscillator and the cannula. Oscillatory signals are analyzed with an on-line computer that measures pressures at points of peak flow. An average resistance is obtained over 8-10 oscillatory cycles. Baseline R^awmeasured in this manner in anesthetized sheep typically results in a value of 1.0-2.0 cm H²O [middle dot] l^-1[middle dot] s^-1, which is close to values reported by others. [29,30] 

Intrabronchial Artery Infusion. Airways reactivity was determined by measuring R^awbefore and after intrabronchial artery infusion of methacholine. Methacholine was delivered through a sideport of the bronchial artery perfusion circuit. From previous experiments, we have confirmed that a plateau in the increase in R^awis achieved within 2 min of agonist delivery. Sheep received a continuous infusion of methacholine in a concentration of 1-2 [micro sign]g/ml at 2 ml/min through the bronchial artery, which caused an [almost equal to] 100% increase in R^aw. With a nominal bronchial artery perfusion rate of 20 ml/min, this delivery rate resulted in calculated molar concentration between 5 x 10^-7m to 10^-6m methacholine. After a 2-min delivery, the infusion pump was turned off and the animal allowed to recover to prechallenge level.

Vagal Nerve Stimulation. The vagus nerves were isolated, and nerve stimulator electrodes were attached bilaterally (Harvard Apparatus, Holliston, MA). After establishing baseline R^aw, the vagal nerves were simultaneously stimulated bilaterally (30 Hz, 30 ms duration, 40 V, 9 s), which caused bronchoconstriction and a decrease in heart rate. Both of these responses rapidly reversed on cessation of stimulation (<30 s).

The sheep were anesthetized and ventilated as described earlier. After a 30-min recovery period (and 2 h after the intramusculary administered ketamine), baseline R^awwas measured, and the airways were constricted first by vagal nerve stimulation (VNS) as described while R^awwas measured. After recovery to baseline (2-3 min), methacholine was infused through the bronchial artery and R^awwas measured again. After recovery to baseline (3-5 min), in random order, the three anesthetic agents were infused into the bronchial artery. The concentration for all the drugs was 5 mg/ml, and the infusion rates were 0.06, 0.20, and 0.60 ml/min. After 10 min of infusion at a each rate, the R^awwas measured prechallenge and during constriction by VNS and infusion of methacholine. After recovery, the next rate was infused and the airway measurements repeated. After the final rate of infusion for a specific drug, the sheep were allowed to recover (30-60 min), baseline measurements were repeated, and the next drug was infused.

The concentration of anesthetic drug in the bronchial circulation was calculated. With a controlled infusion of autologous blood into the bronchial artery at 20 ml/min, and the infusion rates of 0.06, 0.20, and 0.60 ml/min of anesthetic drugs into the perfusate, we calculated the molar concentrations of thiopental to be 5.6 x 10^-5M, 1.9 x 10^-4M, and 5.6 x 10^-4M, respectively. Likewise for propofol, we calculated the molar concentrations to be 8.4 x 10^-5M, 2.8 x 10^-4M, and 8.4 x 10 (^-4), respectively. For ketamine, the calculated molar concentrations were 5.4 x 10^-5M, 1.8 x 10^-4M, and 5.4 x 10.0^-4M, respectively.

Systemic blood pressure was analyzed by one-way analysis of variance. Baseline stimulation (100%) for each sheep for each drug was defined as the change in R^awwith VNS and methacholine before infusion of that specific anesthetic drug into the bronchial artery. The changes in R (^aw) as a percent of baseline stimulation were analyzed separately for each drug by one-way analysis of variance, with Bonferroni correction for repeated measures within the sheep. The effective dose that caused a 50% decrease in baseline response (ED⁵⁰) was calculated along the linear part of the dose-response curves (first dose to third dose) for ketamine and propofol for the VNS and methacholine challenge each sheep. The means of the ED⁵⁰values were compared for each challenge by paired t test. Statistical significance was considered to be P <or= to 0.05.

Baseline systemic blood pressure was 119 +/− 15/88 +/− 16 (systolic/diastolic mean +/− SD) and did not vary significantly during challenges either by drug (P = 0.92) or by dose (P = 0.38). Baseline R^awwas 1.95 +/− 0.14 cm H²O [middle dot] l^-1[middle dot] s^-1. Infusion of the three anesthetic agents into the bronchial artery did not significantly alter the baseline R^awbefore each challenge either by dose (P = 0.88) or by drug (P = 0.83)(Table 1). Further, before infusion of anesthetic drug, VNS and methacholine caused a significant increase in R (^aw) at baseline (maximum response). Vagal nerve stimulation at baseline increased R^awto 5.61 +/− 0.53 cm H²O [middle dot] l^-1[middle dot] s^-1(mean +/− SEM), which was not significantly different among drugs (P = 0.93). Methacholine increased R^awto 3.46 +/− 0.18 cm H (²) O [middle dot] l^-1[middle dot] s^-1, which also did not differ among drugs (P = 0.59).

Thiopental, at all of the doses administered, did not attenuate R (^aw) during either VNS or infusion of methacholine. At concentrations of 5.6 x 10^-5M, 1.9 x 10^-4M, and 5.6 x 10^-4M of thiopental, VNS increased R^awto 94 +/− 25%, 91 +/− 17%, and 80 +/− 28% of control stimulation, respectively (P = 0.92). Similarly, thiopental had no effect on the increase in R^awwith methacholine challenge. Airways resistance increased to 95 +/− 12%, 88 +/− 18%, and 195 +/− 90%, respectively (P = 0.14).

Alternatively, propofol and ketamine had a profound effect on the airway responses to stimulation. Propofol caused a dose-dependent attenuation in the VNS-induced bronchoconstriction. At concentrations of 8.4 x 10^-5M, 2.8 x 10^-4M, and 8.4 x 10^-4M, VNS increased R^awto only 83 +/− 5%, 50 +/− 5%, and 26 +/− 11% of maximum (Figure 1, P < 0.0001). Further, propofol had an effect on methacholine-induced airway constriction but only at the highest concentration. At the concentrations administered, methacholine increased R^awto 124 +/− 19%, 96 +/− 14%, and 43 +/− 27% of maximum (Figure 2, P = 0.05).

Ketamine showed the greatest decrease in the airway response to VNS. At concentrations of 5.4 x 10^-5M, 1.8 x 10^-4M, and 5.4 x 10 (^-4) M, VNS increased R^awto only 87 +/− 19%, 38 +/− 7%, and 8 +/− 2%, respectively (Figure 1, P = 0.0004). At the concentrations delivered, methacholine increased R^awto 114 +/− 14%, 108 +/− 17%, and 56 +/− 17% of maximum (Figure 2, P = 0.14).

For the VNS challenge, the mean ED⁵⁰values for ketamine and propofol were 1.52 +/− 0.58 x 10^-4and 3.54 +/− 0.63 x 10^-4, respectively. The ED⁵⁰value for ketamine was significantly lower than the ED⁵⁰value for propofol during VNS (P = 0.03). For the methacholine challenge, the ED⁵⁰values for ketamine and propofol were 7.93 +/− 3.3 x 10^-4and 5.30 +/− 0.88 x 10^-4, respectively, which were not significantly different (P = 0.38).

Our results show that propofol and ketamine protect against induced airway constriction compared with thiopental. Further, the major mechanism of this bronchoprotection was attenuation of neurally mediated constriction with minimal effects through attenuation of direct airway smooth muscle contraction.

Because the animals needed to be anesthetized during the study, we used a continuous infusion of pentobarbital to maintain anesthesia. We chose pentobarbital because it should not have significant effects on airway reactivity at maintenance doses. [31]In addition, a continuous infusion was used to maintain a constant depth of anesthesia. Because the anesthetic drug challenges were randomized, any changes in depth of anesthesia over time would also be random and would not have biased our results. Further, beyond an adequate depth of anesthesia, deepening barbiturate anesthesia does not appear to influence airway reactivity or tone. [32,33]The finding that the infusion of thiobarbiturate in combination with the pentobarbital intravenous anesthetic agent had no effect on either VNS or methacholine-induced airway constriction also supports the lack of effect of the maintenance pentobarbital anesthesia.

We chose concentrations of drug that would be clinically relevant. In a recent study, Ludbrook et al. [34]examined the rate of administration of propofol on peak arterial concentrations of propofol. When 100 mg of propofol was administered at 200 mg/min, a peak brain arterial concentration of 30 [micro sign]g/ml was measured, which would correspond to a concentration of 1.7 x 10^-4M, and in the middle of our dose range. Therefore, the doses we used appear to be clinically relevant as measured by doses for induction of anesthesia in sheep.

One of our goals was to study the direct bronchoprotective effects of these anesthetic agents and to eliminate any potential confounding effects that these agents might induce through circulating catecholamines systemically. We continuously measured the blood pressure and heart rate in each animal throughout the study. Because the heart rate was profoundly affected by the VNS challenges, we did not analyze this variable as a measure of systemic catecholamine release. In addition, we believed that any increase in systemic catecholamines from the administration of ketamine would be detected easily by increased blood pressure, which we measured continuously by an indwelling arterial catheter. We found no significant changes in blood pressure during the infusion of ketamine nor the other two anesthetic agents into the bronchial artery, even at the highest concentrations. This supports our belief of a lack of significant systemic delivery of the anesthetic agents that were infused into the bronchial artery. Therefore, the decrease in airway responses we observed were local to the airways and not attributable to changes in circulating catecholamines or systemic changes.

Although the effects of inhalational anesthetic agents on baseline airway tone have been demonstrated clearly to cause relaxation, [8]the effects of intravenous agents such as propofol and ketamine are inconclusive. Several investigators have reported relaxant effects of ketamine and propofol on airway tone in vitro, [17,20]and others have reported no effect of these drugs on smooth muscle tone. [18,35]In an older clinical study reported by Huber et al., [14]intravenously administered ketamine caused a dose-dependent decrease in R^awin healthy subjects and in those with acute and chronic reactive airways disease. These patients were intubated, however, which would have increased R^aw. Further, prevention of reuptake of circulating catecholamines from the intravenous administration of ketamine [15]is the most likely explanation of the observed decrease in R^awwith increasing ketamine doses. [23,25]Our results do not support an effect of these drugs on baseline airway tone. We observed no decrease in baseline tone even at the highest concentration delivered directly to the airways. Further, using systemic blood pressure as a marker for increased circulating catecholamines, no change was detected. Therefore, unlike inhalational anesthetic agents, decreased baseline airway tone is unlikely to be an important clinical cause of bronchoprotection by these two agents in asthmatic patients.

The effects of propofol and ketamine at preventing induced bronchoconstriction have been examined more extensively. In vitro [16-21,35,36]and in vivo studies in animals [37]and humans [38-40]have shown that propofol and ketamine are able to attenuate the response to a variety of bronchoconstrictor agents. Consistent with these previous studies, our results also show that propofol and ketamine but not thiopental were able to attenuate induced airway constriction. We found that ketamine and propofol reduced the vagal-induced increase in R^awin a dose-dependent fashion. Although we did not observe complete prevention of the vagal-induced increase in R^aw, this may be attributable to the doses administered or to protein binding. We chose to administer doses that would be achieved clinically during induction of anesthesia. [41-43] 

It is noteworthy that neither drug prevented the methacholine-induced increase in R^aw. Propofol decreased the methacholine-induced bronchoconstriction to 43% of maximum whereas ketamine decreased it to 56% of maximum. The decrease in methacholine-induced bronchoconstriction by propofol did achieve statistical significance (P = 0.05), but that of ketamine did not (P = 0.14). One reason for this marginal statistical significance was attributable to the variability among sheep. Clearly, a decrease to approximately one half in the response to methacholine should be significant. The difference may be accounted for by the slightly different concentrations of drug administered. Although we infused the ketamine and propofol at the same rate, the difference in molecular weight led to a slightly higher molar concentration of propofol to be administered compared with ketamine. Whether reaching statistical significance at the highest dose we infused or at higher does has clinical relevance remains in doubt, however. It is clear that at the lower doses we administered that are clinically relevant, the major effect of these drugs was on neural responses.

Consistent with our findings, several investigators have examined the mechanisms for neural depression by ketamine and propofol. Shrivastav [26]showed that ketamine, applied externally to giant squid axon, depolarized the nerve in a concentration-dependent fashion, reduced inward peak transient currents, and reduced steady-state current. Cronnelly et al. [22]demonstrated that ketamine affected the amplitude but not the frequency of miniature end-plate potentials of frog sartorius muscle.

Further, McGrath et al. [24]showed that ketamine depressed preganglionic sympathetic discharge in a dose-related fashion in rabbits. The results from Lundy and Frew [23]and Nedergaard [25]suggested that ketamine affected neural transmission by blocking extraneuronal uptake of catecholamines through inhibition of a neuronal membrane pump, which transports norepinephrine into the adrenergic neurones. Biddle et al. [44]examined the effects of propofol on the neural responses in a rat artery smooth muscle preparation. They found that propofol attenuated the response to exogenous norepinephrine and the response to endogenous norepinephrine release from nerve terminals induced by electrical field stimulation. Any direct effect of the drugs on smooth muscle, however, would also inhibit a neurally mediated bronchoconstriction. Our findings are consistent with the ability of these drugs to diminish neural responses through prejunctional effects. It was somewhat surprising that we did not observe a decrease in baseline tone; however, this may be related to the resting tone in the sheep.

That the primary mechanism of propofol and ketamine inhibition of bronchoconstriction is through neural mechanisms is also consistent with clinical investigations. Ketamine and propofol have been shown to protect against bronchoconstriction on induction of anesthesia and intubation of the trachea. [10-12]The increase in R^awwith airway manipulation such as bronchoscopy or tracheal intubation is mediated through neural mechanisms, which can also be blocked by the administration of local anesthetic agents. [45]Whether the exact mechanism of neural depression by propofol and ketamine is the same as that of local anesthetic agents remains to be determined.

Finally, whether propofol and ketamine are effective at reversing bronchoconstriction is currently not clear. There is some anecdotal evidence that propofol [40,46]and ketamine [13]can reverse bronchoconstriction. When bronchoconstriction was induced in healthy subjects with ultrasonic aerosols, however, inhaled halothane but not intravenously administered ketamine reversed the increased R^aw. [47]Unfortunately, our study was not designed to address this question.

Propofol and ketamine attenuate induced bronchoconstriction. Both have local effects on the airways, with their major mechanism of bronchoprotection occurring through depression of neurally induced bronchoconstriction. In addition, these drugs depress direct airway smooth muscle activation, but this appears to be less important at clinically relevant concentrations. Furthermore, ketamine is more potent than propofol at preventing neurally induced bronchoconstriction.