Cannabinoid antagonist AM 281 reduces mortality rate and neurologic dysfunction after cecal ligation and puncture in rats.
OBJECTIVE
The purpose of this study was to examine whether anandamide, an endogenous cannabinoid receptor ligand, is involved in the pathogenesis of septic encephalopathy.


METHODS
Prospective, controlled study.


METHODS
Male Wistar rats (7 wks old) were randomly divided into four groups as follows: group 1, control (0.5 mL of saline injected subcutaneously); group 2, sham (surgical abdominal incision and suturing were performed, but ligation and puncture of the cecum were omitted); group 3, cecal ligation and puncture (CLP); group 4, CLP + AM 281 ([N-morpholin-4-yl]-5-[2,4-yl]-5-[2,4-dichlorophenyl]-4-methyl-1H-pyrazole-3-carboxamide) as the cannabinoid receptor antagonist (1 mg/kg intraperitoneally).


METHODS
Sepsis was induced by CLP under pentobarbital anesthesia (10 mg/kg intraperitoneally) with 1% isoflurane. A 2-Fr high-fidelity micromanometer catheter was inserted into the left ventricle via the right carotid artery to assess hemodynamics. Each of the rats was neurologically assessed at 30 mins and 12, 24, and 48 hrs after the treatment. The cytoplasmic levels of caspase-3 in the hippocampi were assayed before surgery and at 30 mins and 24 and 48 hrs after surgery using Western blotting techniques. To examine the effects of AM 281 on neurologic function and mortality rate, we set another control group treated solely with AM 281. Selective inducible nitric oxide synthase inhibitor, L-N6-(1-iminoethyl)-lysine (4 mg/kg), was injected intraperitoneally immediately after CLP to produce the CLP + L-N6-(1-iminoethyl)-lysine group to exclude the influence of depressed hemodynamics on neurologic impairment.


RESULTS
It was found that administration of AM 281 could prevent the hemodynamic changes induced by sepsis. Reflex responses, including the pinna, corneal, paw or tail flexion, and righting reflexes, and the escape response significantly decreased in the CLP and CLP + L-N6-(1-iminoethyl)-lysine groups at 48 hrs after the surgery. In contrast, no changes in these reflex responses were found between the CLP + AM 281 and control and sham groups. In addition, no effects of the administration of AM 281 on neurologic function and mortality rate in the control group were found. Tissue caspase-3 levels were elevated at 48 hrs after CLP in the CLP alone group (means +/- sd: control, 3.9 +/- 0.4; sham, 4.2 +/- 0.4; CLP, 7.1 +/- 1.0 [p < .01]; CLP + AM 281, 4.0 +/- 0.5 densitometric units). In addition, administration of AM 281 also decreased the mortality rate (p < .05).


CONCLUSIONS
Administration of AM 281 prevented the hemodynamic changes and development of neurologic dysfunction occurring in association with septic shock, and could decrease the mortality rate in experimentally induced septic shock in rats. Although further studies are necessary to determine whether endogenous cannabinoids cause septic encephalopathy in rats directly or via their effects on systemic hemodynamics, the beneficial effects of AM 281 on these rats might have significant therapeutic implications in cases of septic encephalopathy.
Kadoi
Yuji
Y
Department of Anesthesiology, Gunma University Graduate School of Medicine, Gunma, Japan.
Hinohara
Hiroshi
H
Kunimoto
Fumio
F
Saito
Shigeru
S
Goto
Fumio
F
D000818:Animals; D001095:Arachidonic Acids / Q000037:antagonists & inhibitors*; D001794:Blood Pressure / Q000187:drug effects; D001930:Brain Injuries / Q000209:etiology / Q000517:prevention & control*; D063385:Cannabinoid Receptor Modulators / Q000037:antagonists & inhibitors*; D002560:Cerebrovascular Circulation / Q000187:drug effects; D003251:Constriction, Pathologic; D063388:Endocannabinoids; D006339:Heart Rate / Q000187:drug effects; D008297:Male; D009025:Morpholines / Q000627:therapeutic use*; D053284:Polyunsaturated Alkamides; D011720:Pyrazoles / Q000627:therapeutic use*; D051381:Rats; D017208:Rats, Wistar; D018805:Sepsis / Q000209:etiology / Q000517:prevention & control*
D001095:Arachidonic Acids; D063385:Cannabinoid Receptor Modulators; D063388:Endocannabinoids; D009025:Morpholines; D053284:Polyunsaturated Alkamides; D011720:Pyrazoles; C109925:AM 281; C078814:anandamide