Coronary blood flow (CBF) is normally matched to myocardial oxygen demand by metabolic adjustments in coronary vasomotor tone [1] 2 ) essentially constant. Previous in vivo studies demonstrated well-maintained CBF during inhalation of enflurane, even though myocardial oxygen demand was reduced, resulting in a decrease in myocardial oxygen extraction [2-4] [1] [2-4] [5,6]
The present study evaluated the direct coronary vasomotor effects of enflurane in vivo under stable hemodynamic conditions. A constant-pressure, extracorporeal perfusion system was used to administer clinically relevant concentrations of enflurane selectively into the left anterior descending coronary artery (LAD) of in situ canine hearts, while measurements of CBF, myocardial oxygen consumption (MVO2 ), and myocardial segmental shortening (SS) were obtained. The new findings during enflurane were compared with previous findings obtained in the same preparation during isoflurane and halothane [7]
Methods
This study was conducted after approval from the institutional research committee. Experiments were performed on 11 mongrel dogs of either sex (weight range 20.5-28.0 kg). Anesthesia was induced with an intravenous (IV) bolus injection of thiopental 15 mg/kg and maintained by continuous IV infusion of fentanyl and midazolam at rates of 12 micro gram centered dot kg-1 centered dot h-1 and 0.6 micro gram centered dot kg-1 centered dot h-1 , respectively. Adequacy of this anesthesia regimen was demonstrated by the lack of muscle movement and of hemodynamic responses during surgical preparation. After tracheal intubation, the lungs were mechanically ventilated (Air Shields, Inc., Hatboro, PA) with the fraction of expired oxygen equal to 1.0. The volume and rate of the ventilation were established to maintain arterial PCO2 at a physiological level (30-40 mm Hg). PO2 , PCO2 , and pH of coronary arterial and venous blood samples (see below) were measured electrometrically (Blood Gas Analyzer model 413; Instrumentation Laboratories, Lexington, MS). Muscle paralysis was obtained with an IV injection of vecuronium bromide 0.1 mg/kg, with supplements at 0.05 micro gram centered dot kg-1 centered dot h-1 , to facilitate mechanical ventilation. Body temperature was maintained at 38 degrees C with a heating pad. Lactated Ringer's solution was administered continuously at a rate of 5 mL centered dot kg-1 centered dot h-1 IV to compensate for evaporative fluid losses. Heparin (400 U/kg with supplementation) was used for anticoagulation.
After a left thoracotomy in the fourth intercostal space, the LAD was cannulated and perfused via an extracorporeal system, as described in detail previously [8] 2-5 ).0% CO2 gas mixture, which passed through a calibrated Fortec vaporizer (Cyprane, Yorkshire, England) providing enflurane. Blood was recirculated at least 15 min through the extracorporeal oxygenator to ensure complete equilibration at the desired enflurane concentration.
The LAD perfusion line was equipped with 1) a heat exchanger to maintain the temperature of coronary perfusate at 38 degrees C, 2) a Doppler flow transducer (Transonic System Inc., Ithaca, NY) to measure CBF, and 3) a port for collecting samples of coronary perfusate. Coronary perfusion pressure (CPP) was measured through a small-diameter tube positioned at the orifice of the perfusion cannula.
Measurements of aortic, left atrial and left ventricular pressures, left ventricular dP/dtmax , and heart rate were obtained using standard methods [8]
Measurements of MVO2 were obtained in the LAD bed by applying the Fick principle. The anterior interventricular vein was cannulated in a retrograde direction to obtain samples of local venous effluent [9,10] 2 ) difference. CBF was constant during the period of blood sampling, which satisfied the requirement of the Fick principle for steady-state conditions. Hemoglobin concentration and percent hemoglobin saturation of these samples were measured with a CO-Oximeter (model 482; Instrumentation Laboratories), and used to calculate O2 bound to hemoglobin, assuming an O2 carrying capacity for hemoglobin of 1.39 mL O2/g . The O2 dissolved in the blood was computed (O2 dissolved = 0.003 mL O2 centered dot 100 mL blood-1 centered dot mm Hg-1 ) and added to the bound component to calculate total O2 content. MVO2 (in mL centered dot min-1 centered dot 100 g-1 ) was calculated as the product of the coronary arteriovenous O2 difference and CBF at the time that blood samples were taken. Myocardial O2 extraction (in percent) was calculated by dividing arteriovenous O2 difference by arterial O2 content.
Measurements of SS, an index of local myocardial contractility, were obtained by sonomicrometry [11] [12] max ), respectively [13] Equation 1 where EDL = end-diastolic length; and ESL = end-systolic length.
Enflurane concentration in samples of coronary arterial blood was determined using a modification of the gas chromatographic technique of Yamamura et al. [14]
After at least 45 min of recovery from surgical preparation, initial control measurements for CBF, MVO2 , and SS were obtained in seven dogs during perfusion from the enflurane-free blood reservoir with CPP at 80 mm Hg. This level of CPP was maintained throughout the study. Adenosine was then infused into the LAD perfusion tubing at 8 mg/min, which was sufficient to cause steady-state, maximum vasodilation [15] 2 , and SS were repeated. The value for CBF during adenosine infusion was divided by the preadenosine baseline CBF to calculate a value for the coronary vasodilator reserve ratio (CVRR). After at least 30 min for recovery, new control measurements were obtained, and the LAD was switched to the enflurane-equilibrated blood reservoir. Intracoronary administration of enflurane caused increases in CBF, which peaked within 3-5 min. These values for CBF were obtained, along with the associated values for SS and MVO2 . After exposure of the LAD bed to enflurane, perfusion was returned to the enflurane-free blood reservoir and at least 30 min was allowed for recovery. This protocol, with each exposure to enflurane being immediately preceded by a control period, was followed for enflurane concentrations of 1.1%, 2.2%, and 4.4% [equivalent to 0.5, 1.0 and 2.0 minimum alveolar anesthetic concentration [MAC] for dogs [16]
In four dogs, experiments were performed to evaluate whether vasoactive substances originating in the oxygenator contributed to the increase in CBF during enflurane administration, and whether ischemia was the cause for the segmental lengthening during enflurane administration. To address the first issue, while the LAD was being perfused with blood equilibrated with 2.2% enflurane, the vaporizer was turned off so that enflurane concentration in the coronary arterial blood declined progressively. A value for CBF was obtained when enflurane was no longer detected in the coronary arterial blood (15-20 min). To address the second issue, two approaches were taken. First, paired coronary arterial and venous blood samples were obtained before and during intracoronary administration of 2.2% enflurane and analyzed for plasma lactate concentration (Paramax Analytic System; Baxter, Irvine, CA). Myocardial lactate uptake was calculated by substituting the coronary arteriovenous difference for plasma lactate concentration and the plasma flow (determined from the CBF and hematocrit) into the Fick equation. Second, adenosine was used to increase CBF maximally while SS was converted to segmental lengthening by 2.2% enflurane.
At the completion of each experiment, Evans blue dye was injected into the LAD perfusion tubing to delineate the LAD perfusion field. The heart was stopped with KCl, and removed, and the dyed tissue was excised and weighed so that CBF could be expressed on a per 100 g basis. The average weight of the LAD perfusion field was 25 +/- 3 g.
The Student's t-test for paired samples was used to assess the difference of values during enflurane and adenosine relative to their respective control values and to compare the duplicate values for CVRR obtained in each preparation [17] [17]
Results
All concentrations of enflurane caused increases in CBF which, at constant perfusion pressure, indicated proportional decreases in coronary vascular resistance Figure 1 . The magnitude of these increases in CBF was essentially concentration dependent, although the difference in the responses during 1.1% and 2.2% of enflurane did not quite achieve statistical significance. With 4.4% enflurane, the level of CBF was similar to the maximum CBF achieved with adenosine. Table 1 shows that the concentration of enflurane in the LAD blood supply was proportional to the percentage of enflurane provided by the vaporizer.
Figure 1: Changes in coronary blood flow, myocardial oxygen consumption, and myocardial segmental shortening during intracoronary administrations of enflurane or adenosine. Values are mean +/- SE. *P < 0.05, versus control.
Table 1: Systemic Hemodynamic Variables and Coronary Arterial and Venous Blood Values During Intracoronary Administrations of Enflurane or Adenosine
Enflurane consistently reduced both segmental contractile function and MVO2 . While 1.1% enflurane diminished SS, 2.2% and 4.4% enflurane converted SS to segmental lengthening Figure 1 . The concentration-related decreases in MVO (2 ) tended to mirror those in segmental contractile function Figure 1 . The combination of reduced MVO2 and increased CBF caused marked decreases in O2 extraction, and the attendant increases in coronary venous PO2 , O2 saturation, and O2 content Table 1 .
Composition of the coronary arterial blood perfusate remained similar to control under all conditions, except that PO2 was increased modestly during enflurane administration Table 1 .
The fivefold increases in CBF during adenosine were accompanied by no changes in SS or MVO2 Figure 1 . Figure 2 shows that the values for CVRR were similar before and after the multiple administrations of enflurane.
Figure 2: Lack of change in coronary vasodilator reserve ratio after multiple administrations of enflurane. Values are mean +/- SE.
(Table 2 ) shows that changing the vaporizer setting from 2.2% to 0.0% enflurane, while continuing LAD perfusion via the oxygenator-supplied reservoir, caused CBF and SS to return to their preenflurane control values.
Table 2: Turning Off the Vaporizer While Continuing Left Anterior Descending Coronary Artery Perfusion Via the Oxygenator-Supplied Reservoir Caused Coronary Blood Flow (CBF) and Segmental Shortening (SS) to Return to Control Values
(Table 3 ) shows that enflurane caused a moderate reduction in percent of myocardial lactate extraction, but it had no effect on myocardial lactate uptake, and that a maximum increase in CBF with adenosine did not reverse the enflurane-induced segmental lengthening.
Table 3: Changes in Coronary Blood Flow (CBF), Segmental Shortening (SS), and Lactate Metabolism During Intracoronary Enflurane Alone and Combined with a Maximally Dilating Infusion of Adenosine
Preenflurane and preadenosine control values did not differ significantly Figure 1 ; Table 1 .
Discussion
In previous studies using the regional coronary perfusion preparation [18,19]
In our preparation, only the enflurane-equilibrated blood reservoir was supplied via a circuit equipped with an oxygenator. Thus, it was necessary to rule out that the oxygenator itself and/or the recirculation protocol used to equilibrate the blood with enflurane contributed to the observed increases in CBF and decreases in SS. This was accomplished using two protocols. First, we demonstrated in a previous study that blood which was recirculated for 15 min through a membrane oxygenator not supplied with a volatile anesthetic had no effects in the LAD bed [15]
Coronary PaO2 was sufficient under all conditions (at least 300 mm Hg) for essentially complete saturation of hemoglobin. Although the values for PaO2 were higher during enflurane (reflecting more efficient gas exchange in the membrane oxygenator compared to the lungs of the dogs), this increased the amount of oxygen dissolved in the plasma and thus added minimally to the total oxygen content of the blood.
The unchanged values for CVRR at the end of each experiment confirmed that the extended experimental protocol did not diminish the ability of the coronary circulation to respond to enflurane, e.g., due to impaired vascular reactivity and/or tissue edema.
Intracoronary enflurane caused coronary vasodilation, which was essentially concentration-dependent. The extent of coronary vasodilation caused by the highest concentration of enflurane (4.4% corresponding to 2.0 MAC) was impressive, being nearly equivalent to that achievable with adenosine. Since enflurane also caused local decreases in SS and MVO2 , reflecting a direct negative inotropic effect, O2 extraction decreased dramatically. These decreases in O2 extraction indicated an uncoupling of coronary O2 supply from myocardial O2 demands, which is the hallmark of a coronary vasodilating drug [1]
Mechanical forces in the left ventricular wall compress the coronary arteries during systole, causing a physical impediment to blood flow [1] [1] [8]
When the coronary vasodilating effect of enflurane was compared to that demonstrated for halothane and isoflurane in the same model [7] Figure 3 . It is difficult to compare our findings to other in vivo studies assessing the relative coronary vasodilating effects of the volatile anesthetics [2-4]
Figure 3: Comparison of the effects of equianesthetic (1 minimum alveolar anesthetic concentration) concentration of enflurane, halothane, and isoflurane on coronary blood flow and myocardial segmental shortening. Data for halothane and isoflurane were adopted from our previous investigation
[7] . The greater than 100% decrease in segmental shortening by enflurane indicates segmental lengthening. Values are mean +/- SE. *P < 0.05, versus enflurane, using analysis of variance and Student-Newman-Keuls test
[16] .
Although systolic lengthening is usually associated with localized myocardial ischemia, it can occur when a potent negative inotrope, e.g., lidocaine, is selectively administered into a branch of the left main coronary artery [20] [21]
Intracoronary enflurane caused significant cardiac depression; SS was reduced substantially at the lowest concentration of enflurane, and was converted to segmental lengthening as enflurane concentration was increased. The ability of enflurane (and of the volatile anesthetics in general) to cause cardiac depression is well established [2-6,22-24] Figure 3 . This supports findings obtained previously in vivo using other experimental approaches to assess the relative inotropic effects of the volatile anesthetics [2,3,24]
Our baseline values for SS were consistent with those obtained previously in open-chest anesthetized dogs [7,20] [25] 2 .
In assessing the clinical relevance of our SS findings, it is important to consider the special conditions of the experimental preparation. Since enflurane was administered selectively into the LAD, it had no systemic effects. This excluded mechanisms which would be expected to mitigate the negative inotropic action of enflurane, e.g., reduction in cardiac afterload and baroreceptor-mediated arousal of the sympathoadrenal system [24]
In summary, the current study demonstrated that abrupt intracoronary administrations of enflurane caused concentration-dependent coronary vasodilation, accompanied by a negative inotropic effect, in in situ canine hearts. The net effect of inspired enflurane on CBF will depend on the extent this direct vasodilating effect is counteracted by moderating factors, including time-dependent vasculature adaptation, and metabolic factors secondary to reduced cardiac workload.
The authors appreciate the expert technical assistance of Derrick L. Harris, BS, and Nancy C. Manabat, MS.
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