Effect of chronic nNOS inhibition on blood pressure, vasoactivity, and arterial wall structure in Wistar rats Sona Cacanyiova a, * , Frantisek Kristek a , Maria Gerova a , Peter Krenek b , Jan Klimas b a Institute of Normal and Pathological Physiology, Centre of Excellence for Cardiovascular Research, Slovak Academy of Sciences, Sienkiewiczova 1, 813 71 Bratislava, Slovak Republic b Department of Pharmacology and Toxicology, Faculty of Pharmacy, Comenius University, Bratislava, Slovak Republic article info Article history: Received 28 August 2008 Revised 4 March 2009 Available online 20 March 2009 Keywords: Neuronal NO synthase Hypotrophy Vasomotor activity abstract While the unequivocal pattern of endothelial nitric oxide (NO) synthase (eNOS) inhibition in cardiovas- cular control has been recognised, the role of NO produced by neuronal NOS (nNOS) remains unclear. The purpose of the present study was to describe the cardiovascular effects of NO production interference by inhibition of nNOS with 7-nitroindazole (7-NI). Wistar rats (10 weeks old) were used: control and exper- imental rats were administered 7-NI 10 mg/kg b.w./day in drinking water for 6 weeks. Systolic blood pressure (BP) was measured by the tail-cuff plethysmographic method. Isolated thoracic aortas (TAs) were used to study vasomotor activity of the conduit artery in vitro. The BP response of anaesthetised ani- mals was used to follow the cardiovascular-integrated response in vivo. Geometry of the TA was measured after perfusion fixation (120 mm Hg) by light microscopy. Expression of eNOS was measured in the TA by immunoblot analysis. Although 6 weeks of nNOS inhibition did not alter systolic BP, the heart/body weight ratio was decreased. Relaxation of the TA in response to acetylcholine (10 À9 –10 À5 mol/L) was moderately inhibited. However, no difference in the BP hypotensive response after acetylcholine (0.1, 1, 10 lg) was observed. The contraction of TA in response to noradrenaline (10 À10 –10 À5 mol/L), and the BP pressor response to noradrenaline (0.1, 1 lg) was attenuated. The inner diameter of the TA was increased, and the wall thickness, wall cross-sectional area, and wall thick- ness/inner diameter ratio were decreased. The expression of eNOS in the TA was increased. In summary, cardiac and TA wall hypotrophy, underlined by decreased contractile efficiency, were observed. The results suggested that two constitutive forms of NOS (nNOS, eNOS) likely participate in regulation of cardiovascular tone by different mechanisms. Ó 2009 Elsevier Inc. All rights reserved. Introduction Nitric oxide (NO) was found to participate in the control of the cardiovascular system. NO is generated through metabolism of the amino acid arginine by the NO synthase (NOS) enzyme [1]. Three isoforms of NOS were discovered in the cardiovascular system: endothelial (eNOS), neuronal (nNOS), and inducible (iNOS), and it has been suggested that each of has the potential to alter BP [2]. In control conditions eNOS and nNOS are constitutively expressed. Both constitutive NOSs were observed in various cells, however, eNOS is predominantly present in endothelial cells [3]. nNOS was found in neurons, and similarly Bredt et al. [4] described a high level of NO in the central nervous system. In addition, nNOS was found in perivascular (non-adrenergic non-cholinergic, sympa- thetic) nerves [5,6]. However, nNOS was also detected in non-neu- ronal cells including cardiomyocytes, endothelial cells, vascular smooth muscle cells, and skeletal muscle cells [7–9]. Several studies demonstrated a high level of nNOS protein in the kidney, especially in macula densa cells [10–12]. Due to the contribution of eNOS inhibition on NO generated in endothelial cells, we hypothesised that the relatively more specific nNOS inhibitor, 7-nitroindazole (7-NI), could help elucidate the role of NO generated by nNOS in the control of the cardiovascular system. Inhibition of eNOS unequivocally elicits blood pressure (BP) increase, arterial wall hypertrophy, increased contractility, and an inhibition of endothelium-derived relaxation [13–18]. Nevertheless, the involvement and role of NO produced by nNOS in cardiovascular control remains unclear, and the experimental data are controver- sial. In different species, it has been shown that inhibition of nNOS does not increase BP [19–22]. However, Ollerstam et al. [23] and Zag- vazdin et al. [24] described a mild increase in BP after inhibition of nNOS by 7-NI. These disparate findings on BP response after nNOS inhibition and the nature of the cardiovascular responses beg the general question of the cause of this discrepancy. Differences in the animals or inhibitors used, as well as in the method of nNOS inhibitor administration, were not found to be the cause. Thus, we decided to investigate the functional properties of the cardiovascular 1089-8603/$ - see front matter Ó 2009 Elsevier Inc. All rights reserved. doi:10.1016/j.niox.2009.03.002 * Corresponding author. Fax: +421 2 552968516. E-mail address: Sona.cacanyiova@savba.sk (S. Cacanyiova). Nitric Oxide 20 (2009) 304–310 Contents lists available at ScienceDirect Nitric Oxide journal homepage: www.elsevier.com/locate/yniox