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Angiotensin II via AT1 receptors may mediate apoptosis in the cardiac conduction system of rats

U. Vongvatcharanon1, S. Vongvatcharanon2, N. Radenahmad1, P. Kirirat1, P. Intasaro1, P. Sobhon3 and T. Parker4, 1Department of Anatomy, Faculty of Science, Prince of Songkla University, Hat-Yai 90112, Thailand, 2Department of Oral Surgery, Faculty of Dentisty, Prince of Songkla University, Hat-Yai 90112, Thailand, 3Department of Anatomy, Faculty of Science, Mahidol University, Bangkok 10400, Thailand and 4School of Biomedical Science, Nottingham University, Nottingham NG7 2UH, UK.

Apoptosis has been suggested as a possible cause of gradual development of complete heart block and fatal arrhythmias associated with absence of the AV node, sinus, and internodal pathways (James et al, 1996). Studies about apoptosis in the heart by means of cardiomyocyte cell culture have demonstrated that angiotensin II (Ang II ) mediates cardiomyocyte apoptosis via angiotensin II type I receptors (AT1) (Cigola et al, 1997). The transgenic m(Ren-2)27 (TG) rat carries the additional Ren-2 gene, the expression of which results in an increase of heart Ang II (Campbell et al, 1995), thus potentially affecting the cell growth/death equilibrium. This study addresses the question of role of Ang II/AT1 receptors mediated apoptosis in the sinoatrial (SA) and atrioventricular nodes (AV).

Six, male 2 week TG and Hannover Sprague Dawley (SD) rats were anaesthetised by pentobarbitone sodium i.p. injection (100 mg/kg). The hearts were removed and fixed in 10% formaldehyde. Following dehydration and embedding in paraffin, 5 μm serial sections were cut then stained with Masson Trichrome to localize SA and AV nodes. The sections containing SA or AV node were processed for either: (a) calculation of apoptotic nuclei following terminal deoxnucleotidyl transferase nick end labelling of 3'-OH ends using Fluorescein-FragEL™; or (b) immunohistochemical labelling with antibodies to the AT1 receptors prior to confocal scanning laser microscopical analysis. Quantification of AT1 receptors was performed by using Microimage analysis software (Olympus).

Group Apoptotic cells/mm2 AT1 receptors (×103)/mm2
SA AV SA AV
SD 0.0400.07 0.1640.12 1.140.17 7.631.91
TG 0.1400.37* 0.4330.11* 1.670.26* 12.503.97*
Data expressed as mean SD (n=6)
* = significant compared with control (P<0.05) (Independent-Sample T-test)

The table shows that the number of apoptotic cell in both the SA and AV node is significantly greater in the TG compared with the SD (p<0.05). Quantification of AT1 receptors within SA and AV node shows that there were significantly more AT1 receptors in the TG compared with the SD (p<0.05). These data suggest that an elevated level of apoptosis in the TG rat heart compared with the controls could be accounted for by Ren-2 derived Ang II active via AT1 receptors.

Campbell, D.J., Rong, P., Kladis, A., Rees, B., Ganten, D. and Skinner, S.L.(1995) Angiotensin and Bradykinin peptides in the TGR (mRen-2)27 rat. Hypertension, 25, 1014-1020.

Cigola, E., Kajstura, L.B., Meggs, L.G. and Anversa, P. (1997) Angiotensin II activates programmed myocyte cell death in vitro. Experimental Cell Research, 231, 363-371.

James, T.N., Martin, E., Willis, P.W. and Lohr, T.O. (1996) Apoptosis as a possible cause of gradual development of complete heart block and fatal arrhythmias associated with absence of the AV node, sinus, and internodal pathways. Circulation, 93, 1424-1432.