Loss of Sodium-Activated Potassium Channel Slack and FMRP Differentially Affect Social Behavior in Mice.

The sodium-activated potassium channel Slack (Slo2.2) is widely expressed in central and peripheral neurons where it is supposed to shape firing properties important for neuronal excitability. Slack activity is enhanced by interaction with the Fragile-X-Mental-Retardation-Protein (FMRP) and loss of FMRP leads to decreased sodium-activated potassium currents in medial nucleus of the trapezoid body neurons of the Fmr1-knockout (KO) mouse representing a mouse model of the human Fragile-X-Syndrome (FXS) and autism. Autism is a frequent comorbidity of FXS, but it is unclear whether Slack is involved in autistic or related conditions of FXS in vivo. By applying a wide range of behavioral tests, we compared social and autism-related behaviors in Slack- and FMRP-deficient mice. In our hands, as expected, FMRP-deficiency causes autism-related behavioral changes in nesting and in a marble-burying test. In contrast, Slack-deficient males exhibited specific abnormalities in sociability in direct and indirect social interaction tests. Hence, we show for the first time that a proper Slack channel function is mandatory for normal social behavior in mice. Nevertheless, as deficits in social behaviors seem to occur independently from each other in FMRP and Slack null mutants, we conclude that Slack is not involved in the autistic phenotype of FMRP KO mice.

cGMP-Elevating Compounds and Ischemic Conditioning Provide Cardioprotection Against Ischemia and Reperfusion Injury via Cardiomyocyte-Specific BK Channels.

BACKGROUND: The nitric oxide-sensitive guanylyl cyclase/cGMP-dependent protein kinase type I signaling pathway can afford protection against the ischemia/reperfusion injury that occurs during myocardial infarction. Reportedly, voltage and Ca2+-activated K+ channels of the BK type are stimulated by cGMP/cGMP-dependent protein kinase type I, and recent ex vivo studies implicated that increased BK activity favors the survival of the myocardium at ischemia/reperfusion. It remains unclear, however, whether the molecular events downstream of cGMP involve BK channels present in cardiomyocytes or in other cardiac cell types. METHODS: Gene-targeted mice with a cardiomyocyte- or smooth muscle cell-specific deletion of the BK (CMBK or SMBK knockouts) were subjected to the open-chest model of myocardial infarction. Infarct sizes of the conditional mutants were compared with litter-matched controls, global BK knockout, and wild-type mice. Cardiac damage was assessed after mechanical conditioning or pharmacological stimulation of the cGMP pathway and by using direct modulators of BK. Long-term outcome was studied with respect to heart functions and cardiac fibrosis in a chronic myocardial infarction model. RESULTS: Global BK knockouts and CMBK knockouts, in contrast with SMBK knockouts, exhibited significantly larger infarct sizes compared with their respective controls. Ablation of CMBK resulted in higher serum levels of cardiac troponin I and elevated amounts of reactive oxygen species, lower phosphorylated extracellular receptor kinase and phosphorylated AKT levels and an increase in myocardial apoptosis. Moreover, CMBK was required to allow beneficial effects of both nitric oxide-sensitive guanylyl cyclase activation and inhibition of the cGMP-degrading phosphodiesterase-5, ischemic preconditioning, and postconditioning regimens. To this end, after 4 weeks of reperfusion, fibrotic tissue increased and myocardial strain echocardiography was significantly compromised in CMBK-deficient mice. CONCLUSIONS: Lack of CMBK channels renders the heart more susceptible to ischemia/reperfusion injury, whereas the pathological events elicited by ischemia/reperfusion do not involve BK in vascular smooth muscle cells. BK seems to permit the protective effects triggered by cinaciguat, riociguat, and different phosphodiesterase-5 inhibitors and beneficial actions of ischemic preconditioning and ischemic postconditioning by a mechanism stemming primarily from cardiomyocytes. This study establishes mitochondrial CMBK channels as a promising target for limiting acute cardiac damage and adverse long-term events that occur after myocardial infarction.

Amplified pathogenic actions of angiotensin II in cysteine-rich LIM-only protein 4-negative mouse hearts.

LIM domain proteins have been identified as essential modulators of cardiac biology and pathology; however, it is unclear which role the cysteine-rich LIM-only protein (CRP)4 plays in these processes. In studying CRP4 mutant mice, we found that their hearts developed normally, but lack of CRP4 exaggerated multiple parameters of the cardiac stress response to the neurohormone angiotensin II (Ang II). Aiming to dissect the molecular details, we found a link between CRP4 and the cardioprotective cGMP pathway, as well as a multiprotein complex comprising well-known hypertrophy-associated factors. Significant enrichment of the cysteine-rich intestinal protein (CRIP)1 in murine hearts lacking CRP4, as well as severe cardiac defects and premature death of CRIP1 and CRP4 morphant zebrafish embryos, further support the notion that depleting CRP4 is incompatible with a proper cardiac development and function. Together, amplified Ang II signaling identified CRP4 as a novel antiremodeling factor regulated, at least to some extent, by cardiac cGMP.-Straubinger, J., Boldt, K., Kuret, A., Deng, L., Krattenmacher, D., Bork, N., Desch, M., Feil, R., Feil, S., Nemer, M., Ueffing, M., Ruth, P., Just, S., Lukowski, R. Amplified pathogenic actions of angiotensin II in cysteine-rich LIM-only protein 4 negative mouse hearts.

Sildenafil Does Not Prevent Heart Hypertrophy and Fibrosis Induced by Cardiomyocyte Angiotensin II Type 1 Receptor Signaling.

Analyses of several mouse models imply that the phosphodiesterase 5 (PDE5) inhibitor sildenafil (SIL), via increasing cGMP, affords protection against angiotensin II (Ang II)-stimulated cardiac remodeling. However, it is unclear which cell types are involved in these beneficial effects, because Ang II may exert its adverse effects by modulating multiple renovascular and cardiac functions via Ang II type 1 receptors (AT1Rs). To test the hypothesis that SIL/cGMP inhibit cardiac stress provoked by amplified Ang II/AT1R directly in cardiomyocytes (CMs), we studied transgenic mice with CM-specific overexpression of the AT1R under the control of the α-myosin heavy chain promoter (αMHC-AT1R(tg/+)). The extent of cardiac growth was assessed in the absence or presence of SIL and defined by referring changes in heart weight to body weight or tibia length. Hypertrophic marker genes, extracellular matrix-regulating factors, and expression patterns of fibrosis markers were examined in αMHC-AT1R(tg/+) ventricles (with or without SIL) and corroborated by investigating different components of the natriuretic peptide/PDE5/cGMP pathway as well as cardiac functions. cGMP levels in heart lysates and intact CMs were measured by competitive immunoassays and Förster resonance energy transfer. We found higher cardiac and CM cGMP levels and upregulation of the cGMP-dependent protein kinase type I with AT1R overexpression. However, even a prolonged SIL treatment regimen did not limit the progressive CM growth, fibrosis, or decline in cardiac functions in the αMHC-AT1R(tg/+) model, suggesting that SIL does not interfere with the pathogenic actions of amplified AT1R signaling in CMs. Hence, the cardiac/noncardiac cells involved in the cross-talk between SIL-sensitive PDE activity and Ang II/AT1R still need to be identified.

KCNMA1 encoded cardiac BK channels afford protection against ischemia-reperfusion injury.

Mitochondrial potassium channels have been implicated in myocardial protection mediated through pre-/postconditioning. Compounds that open the Ca2+- and voltage-activated potassium channel of big-conductance (BK) have a pre-conditioning-like effect on survival of cardiomyocytes after ischemia/reperfusion injury. Recently, mitochondrial BK channels (mitoBKs) in cardiomyocytes were implicated as infarct-limiting factors that derive directly from the KCNMA1 gene encoding for canonical BKs usually present at the plasma membrane of cells. However, some studies challenged these cardio-protective roles of mitoBKs. Herein, we present electrophysiological evidence for paxilline- and NS11021-sensitive BK-mediated currents of 190 pS conductance in mitoplasts from wild-type but not BK-/- cardiomyocytes. Transmission electron microscopy of BK-/- ventricular muscles fibres showed normal ultra-structures and matrix dimension, but oxidative phosphorylation capacities at normoxia and upon re-oxygenation after anoxia were significantly attenuated in BK-/- permeabilized cardiomyocytes. In the absence of BK, post-anoxic reactive oxygen species (ROS) production from cardiomyocyte mitochondria was elevated indicating that mitoBK fine-tune the oxidative state at hypoxia and re-oxygenation. Because ROS and the capacity of the myocardium for oxidative metabolism are important determinants of cellular survival, we tested BK-/- hearts for their response in an ex-vivo model of ischemia/reperfusion (I/R) injury. Infarct areas, coronary flow and heart rates were not different between wild-type and BK-/- hearts upon I/R injury in the absence of ischemic pre-conditioning (IP), but differed upon IP. While the area of infarction comprised 28±3% of the area at risk in wild-type, it was increased to 58±5% in BK-/- hearts suggesting that BK mediates the beneficial effects of IP. These findings suggest that cardiac BK channels are important for proper oxidative energy supply of cardiomyocytes at normoxia and upon re-oxygenation after prolonged anoxia and that IP might indeed favor survival of the myocardium upon I/R injury in a BK-dependent mode stemming from both mitochondrial post-anoxic ROS modulation and non-mitochondrial localizations.