Article, Resuscitation

Does naloxone alone increase resuscitation rate during cardiopulmonary resuscitation in a rat asphyxia model?

Original Contribution

Does naloxone alone increase resuscitation rate during cardiopulmonary resuscitation in a rat asphyxia model?

Meng-Hua Chen MDa,*, Tang-Wei Liu MDa, Lu Xie DPharmb,

Feng-Qing Song MMa, Tao He MMa

aInstitute of Cardiovascular Diseases, the first affiliated hospital of Guangxi Medical University, Nanning 530027, PR China

bDepartment of Physiology, School of Pre-Clinical Sciences, Guangxi Medical University, Nanning 530021, PR China

Received 4 December 2005; accepted 18 January 2006

Abstract Cardiac arrest was induced with asphyxia to identify if naloxone alone increases resuscitation rate during cardiopulmonary resuscitation in a rat asphyxia model. The animals were randomized into either a saline group (Sal-gro, treated with normal saline 1 ml iv, n = 8), a low-dose naloxone group (treated with naloxone 0.5 mg/kg iv, n = 8), or a high-dose naloxone group (HN-gro, treated with naloxone 1 mg/kg iv, n = 8) in a blinded fashion during resuscitation. At the end of 10 minutes of asphyxia, cardiopulmonary resuscitation was started, and each drug was administered at the same time. The rate of restoration of spontaneous circulation was seen in 1 of 8, 3 of 8, and 7 of 8 animals in the Sal-gro, LN-gro, and HN-gro, respectively. The rate of restoration of spontaneous circulation in HN-gro was significantly higher than that in Sal-gro ( P b .05). Naloxone (1 mg/kg) alone can increase resuscitation rate following Asphyxial cardiac arrest in rats.

D 2006

Introduction

Hughes and colleagues [1,2] first described the presence of endogenous peptides with opiate-like activity in 1975. This finding, coupled with the recognition of clinical similarity between opiate overdose and shock rapidly, led to an increase in research regarding the role of endorphins and their antagonists in the pathophysiology and treatment of shock states. It has been reported that the intravenous administration of naloxone can restore blood pressure in animals and humans during anaphylactic [3], septic, and hemorrhagic shock, probably by antagonism of the cardio- vascular depressant effects of endorphins [4].

* Corresponding author.

Researchers have previously reported the efficacy of intratracheal naloxone in reversing opiate-induced respira- tory depression [5,6]. Even when conventional resuscitated drugs were ineffective, intratracheal naloxone can reverse cardiorespiratory arrest and return the heart rate (HR), blood pressure, and ventilation to normal for a patient who has suffered from severe hemorrhagic hypotension because of the rupture of esophageal varices that developed into cardiorespiratory arrest [7].

Because of the fact that several medications may have been administered at the same time, it is difficult to determine the effect of naloxone during cardiopulmonary resuscitation (CPR). An advantage of using animal models in CPR research is the ability to control differing variables that may be impossible to standardize in clinical trials. The purpose of this study is to identify if naloxone alone

0735-6757/$ – see front matter D 2006 doi:10.1016/j.ajem.2006.01.017

increases resuscitation rate following asphyxial cardiac arrest in rats under the same experimental conditions. The hypothesis is that naloxone will have a beneficial effect on the resuscitation rate during CPR.

Table 2 Time from asphyxia to CA and TOSB

Variables

Time from asphyxia

to CA (s, mean F SD)

Time from asphyxia

to TOSB (s, mean F SD)

Sal-gro

LN-gro HN-gro

261 F 25 264 F 40 277 F 41

52 F 11 48 F 12

44 F 4

Material and methods

This study was approved by the Animal Investigation Committee at our university and was performed in accordance with the National Institutes of Health guidelines for ethical animal research.

Animal preparation

Sprague-Dawley rats of both sexes, weighing 180-220 g, were fasted overnight with the exception of free access to water. The animals were anesthetized by intraperitoneal injection of 1 g/kg urethane, placed in a supine position on a surgical board, and the extremities immobilized. The proximal trachea was surgically exposed in the animals, and a 14-gauge cannula was inserted through a tracheosto- my 10 mm caudal to the larynx. The cannula was advanced for a distance of 1 cm into the trachea and secured by ligature, which was also anchored to the skin.

Through the right external jugular vein, an 18-gauge poly- ethylene catheter was advanced through the superior vena cava into the right atrium. right atrial pressure was measured with reference to the midchest using a high sensitivity pres- sure transducer. Another 18-gauge polyethylene catheter was advanced from the left carotid artery into the thoracic aorta for measurement of aortic pressure using the high sensitivity pressure transducer. The void space of the catheters was filled with physiological salt solution containing 5 IU/ml of bovine heparin. The core temperature was measured through a rectal temperature probe. Conventional lead II electrocardiograms were recorded with subcutaneous needles.

The electrocardiogram (lead II), aortic and right atrial pressures were continuously recorded on a desktop computer via a 4-channel physiological recorder (BL-420 E Bio- systems, The Chengdu Technology & Market Co Ltd, China) for subsequent analyses. The coronary perfusion pressure was calculated as the difference between the minimal diastolic aortic pressure and the simultaneously recorded right atrial pressure.

Experimental protocol

After a 10- to 15-minute equilibration period following surgery, 10 minutes of asphyxial CA was induced with asphyxia by clamping the tracheal tubes. CA was deter- mined by loss of aortic pulsations and mean aortic pressure (MAP) b10 mm Hg together with asystole or pulseless electrical activity.

The animals were prospectively randomized to either a saline group (Sal-gro, treated with normal saline 1 ml iv, n = 8), a low-dose naloxone group (LN-gro, treated with naloxone 0.5 mg/kg iv, n = 8), or a high-dose naloxone group (HN-gro, treated with naloxone 1 mg/kg iv, n = 8) in a blinded fashion during resuscitation after 10 minutes of asphyxial CA. At the end of 10 minutes of asphyxia, CPR was initiated, and each drug was administered at the same time. Drug administration was performed only once at this point in all groups. Ventilation was performed by a volume- controlled small animal ventilator (DH-150, the medical instrument of Zhejiang University, China), with room air at 70 breaths/min and tidal volume adjusted to 6 ml/kg. This ventilation was consistently maintained until spontaneous breathing started or 1 hour after restoration of spontaneous circulation , so as to imitate the scenario of no oxygen available in some circumstance. Manual chest com- pression at a rate of 180 compressions/min with equal com- pression-relaxation duration was always performed by the same investigator, who was blinded to hemodynamic monitor tracings and guided by acoustical audio tones. Compression depth was about 30% of anteroposterior chest diameter at maximal compression. ROSC was defined as the return of supraventricular rhythm with a MAP of z20 mm Hg for a minimum of 5 minutes. Failure to restore spontaneous circulation within 10 minutes resulted in discontinuation of resuscitation efforts.

Table 1 Variables at baseline (mean F SD)

Variables

Sal-gro

LN-gro

HN-gro

Body weight (g)

196 F 12

195 F 13

196 F 11

HR (bpm)

427 F 39

406 F 15

412 F 33

Systolic blood pressure (mm Hg)

144 F 10

140 F 10

135 F 7

Diastolic blood pressure (mm Hg)

93 F 14

84 F 13

92 F 4

MAP (mm Hg)

115 F 13

105 F 13

114 F 8

Center venous pressure (mm Hg)

-1 F 1

-1 F 1

-1 F 1

Body temperature (8C)

37.56 F 0.30

37.34 F 0.21

37.58 F 0.28

ences for variables not normally distributed between the groups. Fisher exact test was performed to compare rate of ROSC and rate of AOSB. A 2-tailed value of P b .05 was considered statistically significant.

Table 3 Comparison of ROSC, time of ROSC, and survival

time among the 3 groups M ( P25,P75)

Time was presented as median (25th, 75th percentile). Time of ROSC

represents the time from CPR to ROSC; survival time represents the time from ROSC to TOSB.

The rate of ROSC in HN-gro was significantly higher than that in Sal- gro ( P b .05). The rate of ROSC in LN-gro was higher than that in Sal- gro but did not indicate a statistical significance. There was no significant difference between LN-gro and HN-gro regarding the rate of ROSC. As for time of ROSC and survival time, there was no significant difference between LN-gro and HN-gro. Sal-gro data cannot be compared with other groups because of the low survival rate.

* P b .05 vs Sal-gro.

Groups

Rats

Rate of

ROSC (%)

Time of

ROSC (s)

Survival

time (h)

Sal-gro

8

1 (12.5)

201

2

LN-gro

8

3 (37.5)

58 (50, 90)

4 (2, 25)

HN-gro

8

7 (87.5)*

60 (55, 65)

3 (1, 27)

postresuscitation care“>Postresuscitation care

Hemodynamics and HR monitoring mentioned above were continued for 1 hour, and mechanical ventilation was continued for 1 hour or less after successful resuscitation depending on the condition of the animal’s respiration. The appearance of spontaneous breathing (AOSB) of the animals was observed closely and recorded immediately by the investigators. AOSB was defined as the return of spontane- ous breathing with more than 5 breaths/min under the circumstances of mechanical ventilation. If spontaneous breathing presented with z40 breaths/min for z5 minutes within 1 hour after ROSC, and blood pressure remained stable or increased gradually, then mechanical ventilation could be withdrawn. After 1 hour of intensified observation, all catheters were removed with the tracheal tube left intact. The wound was then sutured. The animals were subsequently returned to their cages and allowed to recover spontaneously without further interventions. The investigators observed the animals until spontaneous breathing stopped. Survival time was defined as the time from ROSC to termination of spontaneous breathing (TOSB).

Necropsy was routinely performed after death of animals,

including resuscitated and unresuscitated ones. Thoracic and abdominal organs were examined for gross evidence of traumatic injures that followed intraperitoneal injection of anesthesia, airway management, vascular cannulation, or precordial compression, and the position of catheters was documented.

Statistical analysis

Data were presented as mean F standard deviation for approximately normally distributed variables and otherwise as median (25th, 75th percentile). Comparisons between time-based measurements within each group were per- formed with analysis of variance repeated measurements. The Mann-Whitney U test was used to determine differ-

Results

Experimental variables

Before asphyxia, no significant differences were ob- served among the 3 groups with regard to body weight, HR, systolic blood pressure, diastolic blood pressure, MAP, cen- ter venous pressure, and body temperature (Table 1).

Time from asphyxia to CA or to TOSB

Time from the initiation of asphyxia to CA was not significantly different among the 3 groups. Time from the initiation of asphyxia to TOSB was not significantly different among the 3 groups either (Table 2).

Results of CPR in each group

The rate of ROSC in HN-gro was significantly higher than that in Sal-gro ( P b .05) but was not significantly different between LN-gro and Sal-gro. However, the animals treated with naloxone exhibited the tendency of shorter ROSC time and longer survival time. Additionally, there appears to be a relationship between the higher dose of naloxone and the higher rate of ROSC (Table 3).

Changes of respiration within 60 minutes after ROSC among 3 groups

There was no significant difference between LN-gro and HN-gro with regard to time of AOSB and time of ventilator withdrawal. Sal-gro data were excluded because of the low survival rate (Table 4).

CPP changes during resuscitation among 3 groups

CPP in Sal-gro was the lowest, whereas CPP in HN-gro was the highest ( P b .05). There was no significant difference between Sal-gro and LN-gro with regard to CPP during the earliest 10 minutes of the resuscitation phase (Fig. 1). The changes of CPP were consistent with the rate of ROSC among the 3 groups.

Table 4 Comparison of the changes of respiration within 60 minutes after ROSC among the 3 groups (mean F SD)

Groups

ROSC

(num)

Rats of AOSB

Time of AOSB (min)

Time of ventilator withdrawal (min)

Sal-gro

1

1

14

30

LN-gro

3

3

15 F 4

45 F 9

HN-gro

7

6

15 F 5

38 F 10

Time of AOSB represents the time from ROSC to AOSB; num, number; min, minute.

60 250

0 5 10 15 20 25 30 35 40 45 50 55 60

Time (min)

LN-gro HN-gro

CPP progress from CPR

HN-gro LN-gro

Sal-gro

50

200

40

CPP (mmHg)

Heart Rate (beats/min)

30

150

20

10

0

-10

0 0.5

1 1.5 2 2.5 3

3.5 4

5 10

100

50

0

Time (min)

Fig. 1 Changes in CPP during the earliest 10 minutes of resuscitation after 10 minutes of untreated asphyxia. Variables were expressed as mean F SEM. CPP in HN-gro was higher than that in Sal-gro ( P b .05).

Fig. 3 Changes in HR during 60 minutes of monitoring after CPR. Variables were expressed as mean F SD.

Changes in MAP and HR during 60 minutes of monitoring after CPR

There was no significant difference between LN-gro and HN-gro with regard to MAP and HR during 60 minutes of monitoring after CPR (Figs. 2 and 3). Sal-gro data were excluded because of the low survival rate.

Necropsy results

Correct placement of catheters was confirmed in all of the animals. No adverse effects of invasive procedures or other traumatic injures were found.

Discussion

The present study suggests that naloxone (1 mg/kg) alone can increase resuscitation rate during CPR in a rat asphyxia

140

0 5 10 15 20 25 30 35 40 45 50 55 60

Time (min)

LN-gro HN-gro

120

100

MAP (mmHg)

80

60

40

20

0

Fig. 2 Changes in MAP during 60 minutes of monitoring after CPR. Variables were expressed as mean F SD.

model. If saline was regarded as 0 mg/kg of naloxone, it appears as if the Therapeutic effects of naloxone was related to the dose-dependence. A higher dose of naloxone might achieve a higher resuscitation rate. However, doses of naloxone higher than 1 mg/kg were not observed in this study. Further experimentation will be required.

Naloxone is a pure narcotic antagonist. It prevents or reverses the effects of opioids, including respiratory depres- sion, sedation, and hypotension. Naloxone does not possess agonist properties and therefore does not produce respiratory depression, psychotomimetic affects, or pupillary constric- tion. In the absence of opioids, it exhibits essentially no pharmacological activity. Naloxone antagonizes the opioid effects by competitive inhibition at the opioid receptor sites. The indication of naloxone is mainly narcotic overdose or narcotic-induced respiratory and central nervous system depression. It is also used to deal with some kinds of shock because the endogenous opioid system is activated in various shock states. The resulting endogenous opiates depress Cardiovascular function, mainly via central delta-opioid re- ceptors [8,9]. Indeed, plasma beta-endorphin levels correlate well with hemodynamic changes in human septic shock [10] and animal hemorrhagic shock [11]. Thus, naloxone rapidly reverses hemodynamic disturbances and even improves survival by opposing endogenous opiates during shock [12]. It is also generally accepted that hypoxia activates the endogenous opioid system [13,14]. Furthermore, endoge- nous opiates have been shown to be involved in the ventilatory control system [15,16]. During acute hypoxia, naloxone opposes endogenous opiates, increasing sponta- neous ventilation [17,18]. Thus, naloxone can be used to

treat all kinds of respiratory suppression [19,20].

The mechanism of action of naloxone during CPR remains unclear. Prior reports have indicated that hypoxia could cause cerebrospinal fluid or plasma levels of beta-endorphin-like immunoreactivity to increase, which was associated with respiratory difficulties, inhibited the vasoconstriction, and

played a role in the pathophysiology of prolonged infant apnea (near-miss sudden infant death syndrome) [21-24]. It is postulated that asphyxial CA might share the similar reaction in the body and cause a release of endogenous opiates that are partially responsible for respiratory, cardiac, and peripheral vascular depression. Moreover, the response to epinephrine may be attenuated by these substances, but this effect can be reversed by naloxone.

An alternative hypothesis is the involvement of catechol- amines and the autonomic nervous system. Naloxone can promptly stimulate Catecholamine release [25-28], increase Sympathetic nerve activity [29], and significantly elevate HR and blood pressure [30]. Therefore, naloxone alone may improve survival rate in the asphyxial rat model compared with the control group.

CPP is a very important indicator that predicts the possibility of resuscitation. In the present study, the change tendency of CPP appears to be associated well with the dose of naloxone: the higher the dose of naloxone, the higher the CPP and the higher the resuscitation rate (saline can be viewed simply as 0 mg/kg of naloxone). The mechanism for this reaction is presently unclear. Naloxone is generally considered to be a narcotic antagonist devoid of pharmaco- logic activity except for its reversal of opioid (narcotic) effects. However, naloxone in its role as a narcotic antagonist may induce hypertension, pulmonary edema, atrial and ventricular arrhythmias, or CA in certain patients. These ad- verse effects may be because of extreme sympathetic nervous system activity [31] and catecholamine surge at a Short period of time. An extremely high dose or an extremely rapid administration of naloxone is likely to cause these adverse effects. In the case of CA and CPR, it is thought that these adverse effects may provide a beneficial effect and a therapeu- tical mechanism that is sought for during CPR. Because asphyxial CA can be viewed simply as the ultimate respiratory and cardiovascular suppression state, the higher sympathetic nervous system activity and the higher level plasma catechol- amines might help to return spontaneous circulation. Ad- ministration of a large bolus of naloxone might result in a reaction similar to the administration of epinephrine, which might be one of the explanations why naloxone alone can increase resuscitation rate in the asphyxial rat model.

In the present study, a shorter time of ROSC and a longer survival time were seen in LN-gro and HN-gro compared with those in Sal-gro, but the difference had no statistical sig- nificance. A likely explanation for these results may be related to fewer animals participating in the study. All in all, the exact mechanism of naloxone during CPR and whether the animals treated with naloxone have a shorter time of ROSC and longer survival time compared with the control group remain unclear, and further experiments will be necessary.

Conclusion

The present study shows that naloxone alone (1 mg/kg) can increase resuscitation rate following asphyxial CA in

rats under the same experimental conditions. The mecha- nism for this effect is unclear, and further experiments will be necessary.

Acknowledgments

This study was supported by the Guangxi Natural Science Foundation of China (no. 0135038). The authors express their gratitude to the staff of the department of physiology for excellent technical help and expert input.

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