The present work provides experimental evidence showing that inducibility and the modulatory role of cardiac mAChR on rat atrial tachyarrhythmia are influenced by the age of the animal. The stimulation protocol for EFS-induced atrial tachyarrhythmia used in the present work was based on a method previously proposed by Godoy et al. (1999
). The results obtained with this method are qualitatively similar to those obtained by other authors who have studied atrial flutter/fibrillation in vivo
and in vitro
(Alessie et al. 1984
; Euler and Scanlon 1987
; Schuessler et al. 1992
; Inoue et al. 1994
; Watanabe et al. 1996
). However, those authors used direct stimulation whereas we have used EFS to minimise the SS applied to the cardiac tissue (Godoy et al. 2002
). Additionally, in the present study, seeking a more quantitative methodological approach, we related SS to arrhythmia inducibility. By using a similar approach, Kirchhof et al. (1998
) showed the antiarrhythmic and proarrhythmic effects on ventricular fibrillation of amiodarone, procainamide and propafenone. Therefore, the use of SS seems to be suitable for the evaluation of arrhythmia inducibility in diverse experimental conditions. Finally, independent of the stimulation method, these in vivo
and in vitro
studies seem to support the idea that the atrial tachyarrhythmia observed in the present work is probably related to reentrant circuit formation provoked by electrical stimulation of the atrial tissue, and, importantly, might also be mediated by cholinergic mechanisms.
Taking into account that the aforementioned studies were performed only with adult animals, the influence of age was not fully evaluated. In the present work, we obtained results that suggest that mAChR are also involved in atrial tachyarrhythmia in infant, young and mature rats, although the electrophysiological and pharmacological characteristics of this tachyarrhythmia are different in mature rats. For example, we observed that the minimum SS necessary for atrial tachyarrhythmia induction progressively decreased from infants to adults, and increased from adult to mature rats (see Fig. ), suggesting that the arrhythmogenic mechanism(s) in atria might alter during the animal’s development. Such alterations could be, at least in part, due to age-related structural and/or functional changes in cardiac tissue, such as alterations in collagen content and cholinergic mechanisms during the animal’s development (Dhein et al. 2001
; Caulfield and Birdsall 1998
; Brodde et al. 1998
; Brodde and Michel 1999
The minimum SS necessary for atrial tachyarrhythmia induction could be influenced by the atrial mass required to induce tachyarrhythmia and/or the anatomical structure of the atrial tissue and cholinergic mechanisms. In this work, we observed that atrial mass increased with age from infant (8 mg), young (19 mg), adult (46 mg) to mature (50 mg) rats. Although the atrial mass of mature rats was similar to that of adult rats, the minimum SS necessary for tachyarrhythmia induction was higher in mature than in adult rats. These results indicate that other non-electrophysiological factors, including an increase in collagen synthesis and reduced mAChR expression (Brodde et al. 1998
; Brodde and Michel 1999
; Lo et al. 2001
), could be involved in the arrhythmia induction process.
In the present work we also observed that EFS-induced atrial tachyarrhythmia was facilitated or inhibited by micromolar concentrations of agonist (carbachol) or antagonist (atropine) of mAChR in infant, young and adult, but not in mature, rats (Figs. –). The treatment with carbachol—a non hydrolysable analog of ACh—did not significantly change EFS-induced atrial tachyarrhythmia in infant, young and adult rats, whereas it was not possible to induce tachyarrhythmia in the presence of atropine at the same age ranges. However, in mature rats, the minimum SS necessary to induce tachyarrhythmia, compared to control condition, decreased in the presence of carbachol and it was the same in the presence of atropine. Therefore, an mAChR antagonist (atropine) prevented atrial tachyarrhythmia induction at all rat age ranges except in mature rats, whereas carbachol facilitated atrial tachyarrhythmia induction at all ages.
The blockade of cardiac mAChR by atropine increased the NT necessary for attempts at tachyarrhythmia induction, but not the minimum SS necessary for tachyarrhythmia induction, at all ages studied except in mature rats. Atropine was also able to prevent the induction of tachyarrhythmia and to reduce AR (atrial frequency) after tachyarrhythmia at all ages studied except in mature rats. These results suggest that stimulation of mAChR facilitates induction of atrial tachyarrhythmia in infant, young and adult rats. The increment of reentrant circuit formation is a plausible mechanism that would, at least in part, be related to this facilitation. However, regardless of the mechanism of this arrhythmogenic facilitation, the results indicate that it is altered in mature rats.
Some previous results could partially explain the age-related decrease of carbachol and atropine effects observed in the present work. For example, Chevalier et al. (1991
), Dhein et al. (2001
) and Brodde et al. (1998
) showed that the density of cardiac mAChR in rats and humans decrease with age. In addition, Lo et al. (2001
) and Brodde and Leineweber (2004
) showed that expression of mRNA encoding mAChR in rat and human hearts was lower in older than in younger subjects. Therefore, it seems reasonable to conclude that the decreased effects of carbachol and atropine on EFS-induced atrial tachyarrhythmia in mature rats observed in the present work could be related to a reduction of function and/or number of cardiac mAChR with the animal’s age.
In summary, the results obtained in the present work indicate that electrically induced atrial tachyarrhythmia is influenced by age-related changes in cholinergic mechanisms. These changes could be related to a reduction in the function and/or number of mAChR, as well as a reduction in endogenous ACh release (Oberhauser et al. 2001
). In humans, these age-related alterations of cholinergic mechanisms could dramatically interfere with the diagnosis and treatment of cardiac dysfunction, e.g. interfering with the dobutamine-stress echocardiography used to evaluate autonomic function in transplanted hearts, and affecting treatment of sinus bradycardia in patients with excessive vagal tonus. This work therefore raises a word of caution, and suggests that the suitableness of a given treatment for cardiac dysfunction involving interventions affecting cholinergic mechanisms could differ with patient age.