Experimental model of pediatric asphyxial cardiopulmonary arrest in rats

Abstract

Objective: Develop a clinically relevant model of pediatric asphyxial cardiopulmonary arrest in rats.

Design: Prospective interventional study.

Setting: University research laboratory.

Subjects: Postnatal day 16-18 rats (n = 9/group).

Interventions: Anesthetized rats were endotracheally intubated and mechanically ventilated, and vascular catheters were inserted. Vecuronium was administered, and the ventilator was disconnected from the rats for 8 mins, whereupon rats were resuscitated with epinephrine, sodium bicarbonate, and chest compressions until spontaneous circulation returned. Shams underwent all procedures except asphyxia.

Measurements and main results: Asphyxial arrest typically occurred by 1 min after the ventilator was disconnected. Return of spontaneous circulation typically occurred <30 secs after resuscitation. An isoelectric electroencephalograph was observed for 30 mins after asphyxia, and rats remained comatose for 12-24 hrs. Overall survival rate was 85%. Motor function measured using beam balance and inclined plane tests was impaired on days 1 and 2, but recovered by day 3, in rats after asphyxia vs. sham injury (p <.05). Spatial memory acquisition measured using the Morris-water maze on days 7-14 and 28-35 was also impaired in rats after asphyxia vs. sham injury (total latency 379 +/- 28 vs. 501 +/- 40 secs, respectively, p <.05). DNA fragmentation was detected in CA1 hippocampal neurons bilaterally 3-7 days after asphyxia. Neurodegeneration detected using Fluorojade B was seen in bilateral CA1 hippocampi and layer V cortical neurons 3-7 days after asphyxia, with persistent neurodegeneration in CA1 hippocampus detected up to 5 wks after asphyxia. CA1 hippocampal neuron survival after asphyxia was 39-43% (p <.001 vs. sham). Evidence of DNA or cellular injury was not detected in sham rats.

Conclusions: This model of asphyxial cardiopulmonary arrest in postnatal day 17 rats produces many of the clinical manifestations of pediatric hypoxic-ischemic encephalopathy. This model may be useful for the preclinical testing of novel and currently available interventions aimed at improving neurologic outcome in infants and children after cardiopulmonary arrest.

Fig. 1

EKG, BP, and EEG monitoring in a typical PND 17 rat after 8 min asphyxial cardiac arrest. Partial recovery of EEG is seen after a 2 min anesthetic washout. Return of spontaneous circulation occurs here at 22 sec post-resuscitation. Note that the monitor speed was changed at several positions in the tracing.

Fig. 2

Body weight comparison between sham and asphyxiated rats recorded on PNDs 17 - 30 and 45 - 51. *P < 0.05 sham vs. asphyxia.

Fig. 3

Functional deficits seen in PND 16 - 18 rats after 8 min asphyxial cardiac arrest. Profound motor deficits are seen in A. beam balance latency and B. inclined plane fall angle on d 1 and 2 vs. sham. C. Modest deficits in the latency to find the hidden platform in the Morris-water maze are seen on d 7-11 and 28-32, intergroup differences were detected (P < 0.05 sham vs. asphyxia). D. Cumulative latencies for all days of hidden platform testing. Mean ± SEM, n = 9/group, *P < 0.05 vs. sham.

Fig. 4

Selective neuronal cell death in the CA1 region of the hippocampus and layer III cortical neurons after asphyxial cardiac arrest in PND 16 – 18 rats. A and C. Fluorojade B labeling (green) of degenerating neurons in CA1 hippocampus 5 wk after sham-injury (A) or asphyxia (C). Inset, higher magnification; sections were co-labeled with DAPI which stains cell nuclei blue. B and D. H&E staining (upper inset, higher magnficiation) and TUNEL (lower inset; brown nuclear labeling) in CA1 hippocampus 7 d after sham-injury (B) or asphyxia (D). E-G. Degenerating neurons in layer III cortical neurons at 7 d.

Fig. 5

CA1 hippocampal neuron density in PND 16 – 18 rats assessed at 5 wk after recovery from 8 min asphyxial cardiac arrest or sham-injury. Mean ± SEM, n = 9/group, *P < 0.001 vs. sham.