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tone. CST peptides dose-dependently (from 11 to 200 nM) reduce contractility (i.e.
LVP, +LVdP/dt), relaxation (−LVdP/dt), and cardiac work (i.e. RPP), while stimu-
late coronary activity as shown by the increased CP (Angelone et al. 2008 ).
Differently from wild-type CST, P370L-CST only reduces contractility, while
G364S-CST is ineffective on the basal mechanical performance. All CST variants
act against β-adrenergic (ISO)-induced positive inotropic and lusitropic effects,
showing a rank order of potency (for ISO-induced positive inotropism:
WT-CST > G364S-CST > P370L-CST; for ISO-induced positive lusitropism:
G364S-CST > WT-CST > P370L-CST (Angelone et al. 2008 ) (Fig. 2 ). Notably, on
both the isolated and Langendorff perfused rat heart (Angelone et al. 2008 ) and rat
papillary muscle (Bassino et al. 2011 ), CST-evoked major depressive actions are
preceded by an early transient positive inotropic effect that disappears 5 min after
peptide administration. This positive inotropism disappears if H1 histamine recep-
tors are inhibited, thus suggesting a role for these receptors. This is consistent with
previous evidence in rat that activation of myocardial H1 receptors mediates
histamine- dependent positive inotropism (Matsuda et al. 2004 ).
Physio-pharmacologic investigations show that CST-induced negative inotro-
pism and lusitropism involve β2-AR and, with lower affinity, β3-AR, but not β1-AR
and cholinergic receptors (Angelone et al. 2008 ). Interestingly, CST-evoked car-
diodepression recruits pertussis toxin (PTX)-sensitive mechanisms, thus calling for
a role of Gi/o proteins. It is known that, contrary to the Gs-mediated β1-AR stimula-
tory effect on contractility and relaxation, β2-AR elicit negative inotropism and
lusitropism via Gi/o proteins (Xiao et al. 1999 ). Thus, the effects induced by CST
through a β2-AR/Gi/o-dependent inhibitory pathway may limit an early Gs-mediated
stimulation of the mechanical contractile performance. In the heart, activation of the
β2-AR/Gi/o cascade recruits PI3K to elicit negative inotropism (Yano et al. 2007 ).
PI3K is an important component of the CST signal transduction mechanism. In fact,
exposure of the rat heart to inhibitors of this kinase abolishes the effects induced by
the peptide (Angelone et al. 2008 , 2012 ). In the heart, downstream PI3K is the NO/
sGC/cGMP cascade (Abi-Gerges et al. 2001 ). This cascade is involved in the CST-
dependent signaling (Angelone et al. 2008 , 2012 ). In fact, in hearts treated with the
peptide, cGMP significantly increases and this in turn affects two of its major tar-
gets, PKG and phosphodiesterases type 2 (PDE2). Thus, these enzymes may con-
tribute to the CST-dependent depression of myocardial contractility. Via PKG, the
peptide may reduce both L-type Ca2+ current and troponin C affinity for calcium
(Angelone et al. 2008 , 2012 ). Other mediators of the CST-dependent effects are
beta-arrestin and phospholamban (PLN) which undergo S-Nitrosylation in the pres-
ence of the peptide (Angelone et al. 2012 ). Beta-arrestin is implicated in G-protein
coupled receptors desensitization and internalization by clathrin coated vescicles.
When S-nitrosylated, it promotes and accelerates β1-AR desensitization, thus blunt-
ing the effects induced by activation of this adrenoceptor (Ozawa et al. 2008 ). In
line with the counter-adrenergic behaviour of CST, the stimulation of beta-arrestin
desensitization, via NO-dependent protein S-Nitrosylation, may be of relevance to
elicit a more prolonged action of the peptide. This action, in addition to the short-
term effects obtained by recruiting kinases, such as PDE2, highlights the presence
T. Angelone et al.