Role of polyamines and cAMP-dependent mechanisms on 5alpha-dihydrotestosterone-elicited functional effects in isolated right atria of rat.

Androgens produce acute vasodilation of systemic, pulmonary, and coronary arteries in several mammal preparations and increase cardiomyocyte contractility. A decrease of the spontaneous beating of sinoatrial cells has also been described. The aim of this study was to characterize the direct effect of 5alpha-dihydrotestosterone on the spontaneous chronotropism and inotropism in the same preparation as an approach to establish the effect on cardiac output and their mechanism of action. The effects were studied on isolated right atria of Wistar rats placed in an organ bath in Tyrode solution at 37 degrees C and bubbled with carbogen. In male rats, the acute administration of 5alpha-dihydrotestosterone, a nonaromatizable derivate of testosterone, elicited a positive inotropism, which was associated with a negative chronotropism. As reported in the left atria, polyamines and beta-adrenoceptors played a role in 5alpha-dihydrotestosterone-elicited positive inotropism because the effect was antagonized by alpha-difluoromethylornithine, an inhibitor of polyamine synthesis, and atenolol, a beta1-adrenoceptor blocker, but not on the negative effect on chronotropism. The androgen increased the sinoatrial node recovery time, suggesting an effect on the mechanisms of spontaneous diastolic depolarization involved in atria pacemaking. These effects of 5alpha-dihydrotestosterone are not hormonally regulated because they are similarly produced in estrogenized females and gonadectomized male and female rats. These results suggest that the androgen could acutely improve cardiac performance.

Influence of gender and sex hormones on 5alpha-dihydrotestosterone elicited effect in isolated left atria of rats: Role of beta-adrenoceptors and ornithine decarboxylase activity.

Androgens elicit an acute cardiotonic effect in cardiac preparations of rats. This effect is produced via an extracellular interaction that may be coupled to pertussis-sensitive G-proteins and is associated with an increase in cAMP, polyamine synthesis and intracellular calcium. The nature of the targets and the existence of a dimorphic effect in this nongenomic effect of androgens are unknown. The purpose of this study was to characterize a possible gender and sex hormone influence on the 5alpha-dihydrotestosterone-elicited cardiotonic effect, taking into account the possible role of the beta-adrenoceptors and ornithine decarboxylase activity on this response. [Float1]Regarding this, the effect of 5alpha-dihydrotestosterone on isolated left atria from male, estrogenized female and gonadectomized male and female rats was studied. The results showed that 5alpha-dihydrotestosterone-elicited cardiotonic effect was preserved independent of gender and sex hormones, being higher in control males than in the rest of the groups. This correlated with the testosterone plasma levels, except in estrogenized females, suggesting that the androgens positively and the estrogens negatively regulated the response. In all groups, 5alpha-dihydrotestosterone produced an increase in cAMP levels, but only in control males did it produce an increase in ornithine decarboxylase activity. In the other groups, the absence of an effect on ornithine decarboxylase might limit the capability of the response to the androgen. Altogether, androgens may help to control cardiac performance by a direct interaction on the heart in both sexes. Gender and sex differences in the magnitude of inotropism being due mainly to changes in beta-adrenoceptors and cAMP production and in intracellular polyamine synthesis.

Putrescine modulation of acute activation of the beta-adrenergic system in the left atrium of rat.

Endogenous polyamines mediate acute metabolic effects and cardiac hypertrophy associated to beta-adrenoceptor stimulation. The aim of this study is to characterize the role of polyamines on beta-adrenoceptor system mediated responses. To this end, the functional interaction of polyamine modifying drugs on isoproterenol-elicited cardiotonic effect, in isolated left atria of male Wistar rats, and their effects on [(3)H]dihydroalprenolol (DHA) binding on beta-adrenoceptors and on adenylyl cyclase activity of membrane heart were studied. Polyamines interact with beta-adrenoceptors in rat heart, as shown by the displacement of [(3)H]DHA binding. Furthermore, putrescine (but not spermidine or spermine) increased adenylyl cyclase activity, elicited a positive inotropism and increased intracellular cAMP. The putrescine effect on adenylyl cyclase was not antagonized by the beta-adrenoceptors blockers, alprenolol and ICI-118,551, and facilitated the isoproterenol effect. Neither alprenolol, atenolol nor ICI-118,551 antagonized putrescine-elicited positive inotropism. However, the effect was abolished in preparations with desensitized beta-adrenoceptors. alpha-Difluoromethylornithine, an inhibitor of ornithine decarboxylase, antagonized the effect of isoproterenol on inotropism and cAMP increase. In addition, putrescine might elicit effects by mechanisms independent of beta-adrenoceptor system, since in left atria with functional desensitized receptors an interaction with ouabain-elicited cardiotonic effect was observed. These results suggest that putrescine may act as a low affinity agonist on beta-adrenoceptors and modulate acute responses mediated by beta-adrenoceptors. These findings may be of importance in the physiology and in diseases involving cardiac beta-adrenoceptors.

Role of putrescine on androgen-elicited positive inotropism in the left atrium of rats.

Functional and biochemical studies were performed in isolated left atria of male Wistar rats to study whether endogenous polyamines may mediate androgen-elicited positive inotropism and their relationship with a rise in cAMP during the cardiotonic effect. 5 alpha-Dihydrotestosterone (100 microM) exposure increased intracellular putrescine as determined by HPLC, but it did not increase spermidine and spermine. This effect was antagonized by an inhibitor of ornithine decarboxylase, alpha-difluoromethylornithine (10 mM), suggesting enzyme activation. alpha-Difluoromethylornithine also antagonized androgens-elicited inotropism and the increase in intracellular cAMP. Putrescine (1 to 10 mM) elicited a concentration-dependent positive inotropism associated with the cAMP increase. The prior incubation with putrescine antagonized 5 alpha-dihydrotestosterone-elicited inotropism and did not produce sinergism on intracellular cAMP. Short-term incubation with 5 alpha-dihydrotestosterone or forskolin shifted to the left the cardiotonic effect of isoproterenol, an agonist of beta-adrenoceptors, without any increase in Emax, suggesting that a common mechanism was involved. Therefore, polyamines might modulate the cAMP production associated with the cardiotonic effect of androgens.

Role of the C-terminus of the high-conductance calcium-activated potassium channel in channel structure and function.

The role of ion channels in cell physiology is regulated by processes occurring after protein biosynthesis, which are critical for both channel function and targeting of channels to appropriate cell compartments. Here we apply biochemical and electrophysiological methods to investigate the role of the high-conductance, calcium-activated potassium (Maxi-K) channel C-terminal domain in channel tetramerization, association with the beta1 subunit, trafficking of the channel complex to the cell surface, and channel function. No evidence for channel tetramerization, cell surface expression, or function was observed with Maxi-K(1)(-)(323), a construct truncated three residues after the S(6) transmembrane domain. However, Maxi-K(1)(-)(343) and Maxi-K(1)(-)(441) are able to form tetramers and to associate with the beta1 subunit. Maxi-K(1)(-)(343)-beta1 and Maxi-K(1)(-)(441)-beta1 complexes are efficiently targeted to the cell surface and cannot be pharmacologically distinguished from full-length channels in binding experiments but do not form functional channels. Maxi-K(1)(-)(651) forms tetramers and associates with beta1; however, the complex is not present at the cell surface, but is retained intracellularly. Maxi-K(1)(-)(651) surface expression and channel function can be fully rescued after coexpression with its C-terminal complement, Maxi-K(652)(-)(1113). However coexpression of Maxi-K(1)(-)(343) and Maxi-K(1)(-)(441) with their respective C-terminal complements did not rescue channel function. Together, these data demonstrate that the domain(s) in the Maxi-K channel necessary for formation of tetramers, coassembly with the beta1 subunit, and cell surface expression resides within the S(0)-S(6) linker domain of the protein, and that structural constraints within the gating ring in the C-terminal region can regulate trafficking and function of constructs truncated in this region.

Interaction of androgens with cardiotonic drugs in isolated left atrium of rat.

Pharmacological concentrations of androgens are known to elicit a rapid positive inotropism in isolated left atrium of male rats. Upon short-term exposure to androgens, an increase in intracellular cAMP levels has been observed, though delayed with respect to the time course of contraction, suggesting that other mechanisms may participate in initiating the contraction. Therefore, the interaction of positive inotropism elicited by ouabain, an inhibitor of Na(+)-K(+)-ATPase, and androgens was studied in isolated left atrium of rat. Androgens antagonized ouabain-elicited positive inotropism and increased the basal tone. Vanadate, an inhibitor of the Ca(2+) pump, produced a similar effect as androgens on ouabain-elicited positive inotropism. Therefore, androgens might interact with the Ca(2+) pump and this may explain the increase in basal tone. The conjugation of 5 alpha-dihydrotestosterone with bovine serum albumin produced the same effect, suggesting an extracellular interaction of androgens inhibiting the Na(+)-K(+)-ATPase that could increase intracellular Ca(2+) via the Na(+)-Ca(2+) exchange.