Regulation of Papillary Muscle Contractility by NAD and Ammonia Interplay: Contribution of Ion Channels and Exchangers.

Various models, including stem cells derived and isolated cardiomyocytes with overexpressed channels, are utilized to analyze the functional interplay of diverse ion currents involved in cardiac automaticity and excitation-contraction coupling control. Here, we used ß-NAD and ammonia, known hyperpolarizing and depolarizing agents, respectively, and applied inhibitory analysis to reveal the interplay of several ion channels implicated in rat papillary muscle contractility control. We demonstrated that: 4 mM ß-NAD, having no strong impact on resting membrane potential (RMP) and action potential duration (APD90) of ventricular cardiomyocytes, evoked significant suppression of isometric force (F) of paced papillary muscle. Reactive blue 2 restored F to control values, suggesting the involvement of P2Y-receptor-dependent signaling in ß-NAD effects. Meantime, 5 mM NH4Cl did not show any effect on F of papillary muscle but resulted in significant RMP depolarization, APD90 shortening, and a rightward shift of I-V relationship for total steady state currents in cardiomyocytes. Paradoxically, NH4Cl, being added after ß-NAD and having no effect on RMP, APD, and I-V curve, recovered F to the control values, indicating ß-NAD/ammonia antagonism. Blocking of HCN, Kir2.x, and L-type calcium channels, Ca2+-activated K+ channels (SK, IK, and BK), or NCX exchanger reverse mode prevented this effect, indicating consistent cooperation of all currents mediated by these channels and NCX. We suggest that the activation of Kir2.x and HCN channels by extracellular K+, that creates positive and negative feedback, and known ammonia and K+ resemblance, may provide conditions required for the activation of all the chain of channels involved in the interplay. Here, we present a mechanistic model describing an interplay of channels and second messengers, which may explain discovered antagonism of ß-NAD and ammonia on rat papillary muscle contractile activity.

Anti-hypoxic effect of interleukin-10 in hippocampal neurons is mediated by modulation of TASK-1 and TASK-3 channels activity.

It has been shown that anti-inflammatory cytokine interleukin-10 (IL-10) can exert anti-hypoxic effect preventing post-hypoxic neuronal hyperexcitability. Yet, exact mechanisms of IL-10 mediated anti-hypoxic action on neuronal function are not fully understood. We suggested that IL-10 can exert its anti-hypoxic action via modulation of activity of two-pore potassium TASK-1 and TASK-3 channels. To study the involvement of TASK-1 and TASK-3 channels we employed a combination of whole-cell patch clamp and pharmacological inhibitory analysis to assess if IL-10 and brief hypoxic episode can modulate K+ background leak current (Ileak) and membrane input resistance (Rin) in cultured hippocampal neurons. We found that IL-10 in a dose-dependent manner can significantly increase Ileak with concomitant reduction in Rin. Neurons that were exposed to brief hypoxic episode on contrary showed significant decrease in Ileak with concomitant increase in Rin. Pretreatment with IL-10 prior hypoxic episode was able to abolish negative effect of hypoxia on Ileak and Rin. IL-10 potentiating action on Ileak and Rin was occluded by co-addition of selective blockers of TASK-1 and TASK-3 channels - ML365 and PK-THPP. Co-addition of LY294002, an inhibitor of PI3-kinase occluded IL-10 action on Ileak and Rin showing involvement of PI3K-associated pathway in IL-10 mediated regulation of TASK channel function. Our results provide new insights into IL-10 mediated neuroprotective and anti-hypoxic actions showing TASK-1 and TASK-3 channels as downstream targets of this anti-inflammatory cytokine.

Why Do Levels Of Anti-inflammatory Cytokines Increase During Memory Acquisition?

The role of anti-inflammatory cytokines in the mechanisms of learning and memory, modulation of synaptic plasticity in the mammalian brain has not received sufficient attention. These issues are discussed in this review, and among the many cytokines, attention is paid to the most studied in this respect IL-10, IL-4, IL-13 and TGF-ß. The level of anti-inflammatory cytokines in the brain tends to increase during memory acquisition, but the significance of such an increase is unclear. We hypothesize that anti-inflammatory cytokines primarily protect and optimize the functioning of neuronal circuits involved in information processing. The increased local activity of neurons during memory acquisition activates many signaling molecules, and some of them can trigger unwanted processes (including neuroinflammation), but increased levels of anti-inflammatory cytokines prevent this triggering. Each of the anti-inflammatory cytokines plays a specific role in supporting information processing. For example, the role of IL-4 and IL-13 in recruiting T cells to the meninges during training in healthy animals has been most studied. It has also been shown that TGF-ß is able to optimize late stage LTP in the hippocampus and support the consolidation of memory traces in behavioral studies. Cytokines have an effect on learning and memory through their influence on neuroplasticity, neurogenesis in the hippocampus and regulation of the neurovascular unit. Experiments have shown such an effect, and the data obtained create the prerequisites for new therapeutic approaches to the correction of cognitive impairments.

Interleukin-10 and transforming growth factor-ß1 facilitate long-term potentiation in CA1 region of hippocampus.

It has been shown that pro-inflammatory cytokines preferentially attenuate long-term potentiation (LTP), at the same time the effect of anti-inflammatory cytokines on synaptic plasticity has not been fully studied yet. Here we studied the effect of two anti-inflammatory cytokines - interleukin-10 (IL-10) and transforming growth factor-ß1 (TGF-ß1) on long-term potentiation. It was found that exogenously added IL-10 as well as TGF-ß1 were able to effectively facilitate LTP evoked with ether high frequency or theta burst stimulation protocols in CA1 area of hippocampus. Effectiveness of IL-10 and TGF-ß1 on LTP varied depending on the concentration of used cytokine and type of tetanic stimulation protocol used for LTP induction. Overall the positive effect of studied cytokines on LTP was associated with their ability to increase basal synaptic strength at Schaffer collateral - CA1 synapse. At the same time IL-10 and TGF-ß1 did not have any effect on short-term plasticity. Our results provide new evidence upon the modulatory effects that anti-inflammatory cytokines exert on synaptic plasticity further highlighting their potency as modulators of neuronal function.

Interleukin-10 Facilitates Glutamatergic Synaptic Transmission and Homeostatic Plasticity in Cultured Hippocampal Neurons.

Anti-inflammatory cytokines are known to exert neuroprotective action ameliorating aberrant neuronal network activity associated with inflammatory responses. Yet, it is still not fully understood if anti-inflammatory cytokines play a significant role in the regulation of synaptic activity under normal conditions. Thus, the aim of our study was to investigate the effect of Interleukin-10 (IL-10) on neuronal synaptic transmission and plasticity. For this we tested the effect of IL-10 on miniature excitatory postsynaptic currents (mEPSC) and intracellular Ca2+ responses using whole-cell patch clamp and fluorescence microscopy in 13-15 DIV primary hippocampal neuroglial culture. We found that IL-10 significantly potentiated basal glutamatergic excitatory synaptic transmission within 15 min after application. Obtained results revealed a presynaptic nature of the effect, as IL-10 in a dose-dependent manner significantly increased the frequency but not the amplitude of mEPSC. Further, we tested the effect of IL-10 on mEPSC in a model of homeostatic synaptic plasticity (HSP) induced by treatment of primary hippocampal culture with 1 µM of tetrodotoxin (TTX) for a 24 h. It was found that 15 min application of IL-10 at established HSP resulted in enhanced mEPSC frequency, thus partially compensating for a decrease in the mEPSC frequency associated with TTX-induced HSP. Next, we studied if IL-10 can influence induction of HSP. We found that co-incubation of IL-10 with 1 µM of TTX for 24 h induced synaptic scaling, significantly increasing the amplitude of mEPSC and Ca2+ responses to application of the AMPA agonist, 5-Fluorowillardiine, thus facilitating a compensatory postsynaptic mechanism at HSP condition. Our results indicate that IL-10 potentiates synaptic activity in a dose- and time-dependent manner exerting both presynaptic (short-term exposure) and postsynaptic (long-term exposure) action. Obtained results demonstrate involvement of IL-10 in the regulation of basal glutamatergic synaptic transmission and plasticity at normal conditions.