Morphological and Functional Alterations of the Cardiovascular System during Acute Clozapine Poisoning (Experimental Study)

Purpose of the study. To evaluate the influence of sublethal dose of Clozapine on the functional and morphological parameters of the cardiovascular system in rats 4 hrs. after the drug administration.

Materials and methods. The experiments were carried out on male Wistar rats weighing 200–250g (n=14). Group I received 0.9% NaCl solution administered via enteral feeding tubing under general anesthesia using Sevoflurane; group II — Clozapine dosed at 150 mg/kg in 0.9% NaCl solution; group III — Clozapine dosed at 150 mg/kg in 40% ethyl alcohol solution. 4 hrs. after drug administration, arterial blood pressure (ABP), heart rate (HR), microcirculation in the skin using laser Doppler flowmetry (LSF), fluorescence intensity of coenzymes NADH and FAD were evaluated. After euthanasia, autopsy including withdrawal of the internal organs of rats for morphological analysis was performed. Thereafter, paraffin sections of hearts were made and subsequently stained with hematoxylin and eosin, which were later examined with the help of light microscope Nikon Eclipse Ni-U.

Results. In both Clozapine groups, ABP and blood flow in the skin were lower than in the control group: ABP — by 12% in group I and by 15% in group II; blood flow — by 48% and 37%, respectively. No significant difference between the groups was observed in respect of the microcirculation indices studied. Increased fluorescence intensity of coenzyme NADH in the skin was found in the Clozapine groups compared to the control group (2.3-fold in group I and 1.9-fold in group II). Histological analysis of the hearts of animals that received Clozapine established uneven blood filling, erythrocyte sludges, fine-focal hemorrhaging, endotheliocyte nuclei oriented perpendicular to the basal membrane, uneven staining of myocardium with hypereosinophilic segments, fragmentation and undulating deformity of muscle fibers.

Conclusion. During acute Clozapine poisoning, morphological signs of disturbed circulation and myocardium damage are found in the heart, which are accompanied with development of myocardium dysfunction with arterial hypotension and decreased peripheral blood flow, also disturbed oxidative metabolism in peripheral tissues.

1. Green A., Tohen M., Patel J.K., Banov M., DuRand C., Berman I., Chang H., Zarate C., Posener J., Lee H., Dawson R., Richards C., Cole J.O., Schatzberg A.F. Clozapine in the treatment of refractory psychotic mania. Am. J. Psychiatry. 2000; 157 (6): 982–986. DOI: 10.1176/appi.ajp.157.6.982. PMID: 10831480

2. Tochilov V.A., Kushnir O.N. Clozapine is the drug of choice for the treatment of patients with acute psychosis. Socialnaya i klin. psikhiatriya.2011; 21 (2): 37–42.[In Russ.]

3. Shigeev S.V. Ivanova N.A. Ivanov S.V. Clozapine poisoning: theoretical aspects and forensic assessment. Sud.Med.Expert. 2013; 6: 41–46. [In Russ.]

4. Slyundin D.G., Livanov A.S., Anuchin V.V., Merkin A.G., Bobrinskaya I.T., Tutova E.V. Features of psychopathological manifestations in criminal poisoning with clozapine. Nevrologiya, nejropsikhiatriya, psikhosomatika, 2010; 3: 57–63. [In Russ.]

5. Flanagan R.J., Spencer E.P., Morgan P.E., Barnes T.R., Dunk L. Suspected clozapine poisoning in the UK/Eire, 1992—2003. Forens Sci Int. 2005; 155: 2–3: 91–99. DOI: 10.1016/j.forsciint.2004.10.024. PMID: 16226146

6. Slyundin D.G., Livanov A.S., Anuchin V.V., Bobrinskaya I.G., Gutova E.V. Criminal Clozapine poisonings. Anastes. i reanimatol. 2007; 4: 61–64. [In Russ.] PMID:17929492.

7. Khan A.A., Ashraf A., Baker D., Al-Omary M.S., Savage L., Ekmejian A., Singh R.S.H., Brienesse S., Majeed T., Gordon T., Drinkwater V., Collins N.J. Clozapine and incidence of myocarditis and sudden death – long term Australian experience. Int. J. Cardiol. 2017; 238: 136–139. DOI: 10.1016/j.ijcard.2017.03.013. PMID: 28343762

8. Katta N., Balla S., Aggarwal K. Clozapine-induced hypersensitivity myocarditis presenting as sudden cardiac death. Autops. Case Rep. 2016; 6 (4): 9-13. DOI: 10.4322/acr.2016.054. PMID: 28210568

9. Longhi S., Heres S. Clozapine-induced, dilated cardiomyopathy: a case report. BMC Res. Notes. 2017; 10 (1): 338. DOI: 10.1186/s131040172679-5. PMID: 28750684

10. Woloszyn E., Whig N., Trigoboff E., Grace J.J. Cardiac arrest with clozapine and olanzapine: revealing long QT syndrome. Clin. Schizophr. Relat. Psychoses. 2016. [Epub. ahead of print]. DOI: 10.3371/CSRP.WOWH.112316. PMID: 27996317

11. Barry A.R., Windram J.D., Graham M.M. Clozapine-associated myocarditis: case report and literature review. Can. J. Hosp. Pharm. 2015; 68 (5): 427–429. DOI: 10.4212/cjhp.v68i5.1493. PMID: 26478592

12. Abdel-Wahab B.A., Metwally M.E. Clozapine-induced cardiotoxicity: role of oxidative stress, tumour necrosis factor alpha and NF-κβ. Cardiovasc. Toxicol. 2015; 15 (4): 355–365. DOI: 10.1007/s12012-0149304-9. PMID: 25539628

13. Abdel-Wahab B.A., Metwally M.E. Clozapine-induced cardiotoxicity in rats: involvement of tumour necrosis factor alpha, NF-�βand caspase3. Toxicol. Rep. 2014; 1: 1213–1223. DOI: 10.1016/j.toxrep.2014. 11.012. PMID: 28962331

14. Williams D.P., O’Donnell C.J., Maggs J.L., Leeder J.S., Uetrecht J., Pirmohamed M., Park B.K. Bioactivation of clozapine by murine cardiac tissue in vivo and in vitro. Chem. Res. Toxicol. 2003; 16 (10): 1359–1364. DOI: 10.1021/tx034035z. PMID: 14565776

15. Ishiyama S., Hiroe M., Nishikawa T., Abe S., Shimojo T., Ito H., Ozasa S., Yamakawa K., Matsuzaki M., Mohammed M.U., Nakazawa H., Kasajima T., Marumo F. Nitric oxide contributes to the progression of myocardial damage in experimental autoimmune myocarditis in rats. Circulation. 1997; 95 (2): 489–496. DOI: 10.1161/01.CIR.95.2.489. PMID: 9008468

16. Lee S.Y., Kim Y.J., Kim K.T., Choe H., Jo S.H. Blockade of HERG human K+ channels and IKr of guinea-pig cardiomyocytes by the antipsychotic drug clozapine. Br. J. Pharmacol. 2006; 148 (4): 499–509. DOI: 10.1038/sj.bjp.0706744. PMID: 16633353

17. Raheja H., Namana V., Chopra K., Sinha A., Gupta S.S., Kamholz S., Moskovits N., Shani J., Hollander G. Electrocardiogram Changes with Acute Alcohol Intoxication: A Systematic Review. Open Cardiovasc Med J. 2018; 12: 1–6. DOI: 10.2174/1874192401812010001. PMID: 29541259

18. Zoroastrov O.M. Features of thanatogenesis upon death from acute ethanol intoxication. Vestn. Sud. Med. 2016; 3: 42–44. [In Russ.]

19. Directive 2010/63 / EU of the European Parliament and the Council of the European Union on the Protection of Animals used for scientific purposes. St. Petersburg: Rus-LASA NP «Association of Specialists Working with Laboratory Animals» Working Group on translations and publication of subject literature; 2012: 48.[In Russ.]

20. Ryzhkov I.A., Zarzhecky Yu.V. Effect of circulatory and hemic hypoxia on microcirculation in the skin. Regionarnoe krovoobrashchenie i mikrotsirkulyatsiya. 2018; 17(2): 64–70. [In Russ.] DOI: 10.24884/1682-6655-2018-17-2-64-70

21. Ryzhkov I.A., Zarzhetsky Yu.V., Novoderzhkina I.S. Comparative Aspects of the Regulation of Cutaneous and Cerebral Microcirculation During Acute Blood Loss. Obshchaya reanimatologiya=General Reanimatology. 2017; 13 (6): 18–27. [In Russ., In Engl.] DOI: 10.15360/1813-9779-2017-6-18-27

22. Mayevsky A., Rogatsky G.G. Mitochondrial function in vivo evaluated by NADH fluorescence: from animal models to human studies. Am J Physiol Cell Physiol. 2007; 292 (2): 615–640. PMID: 16943239. DOI: 10.1152/ajpcell.00249.2006

23. Krupatkin A.I., Sidorov V.V.Functional diagnostics of the state of microcirculatorytissue systems: Oscillations, information, nonlinearity (A guide for physicians). M .: Book house «LIBROCOM»; 2013: 496. [In Russ.]ISBN 978-5-9710-3329-5