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Modulation of cytosolic-[Ca2+] oscillations in hepatocytes results from cross-talk among second messengers. The synergism between the alpha 1-adrenergic response, glucagon and cyclic AMP, and their antagonism by insulin and diacylglycerol manifest themselves in the control of the cytosolic-[Ca2+] oscillations.

ABSTRACT: Hepatocytes respond to stimulation by glycogenolytic agonists acting via phosphoinositide (PI) breakdown through oscillations of the free cytosolic concentration of Ca2+ ([Ca2+]cyt.). Since the second-messenger repertoire of hepatocytes includes many other factors besides Ca2+, we investigated to what degree the regulation of [Ca2+]cyt. oscillations is integrated into these other signalling systems. [Ca2+]cyt. was recorded in single rat hepatocytes by using the Ca(2+)-indicator fura-2. Parallel stimulation with phenylephrine (an alpha 1-adrenergic agonist of PI breakdown) and glucagon resulted in a synergistic stimulation of [Ca2+]cyt. oscillations. Direct activation of the cyclic-AMP-dependent pathway with several stimuli (forskolin, 8-bromo cyclic AMP, 8-CPT cyclic AMP) mimicked the response to glucagon. In contrast, [Ca2+]cyt. oscillations induced by various combinations of these agonists could be antagonized by the glycogenic hormone insulin. As one of the options in the insulin-signalling network, we tested a diacylglycerol activator of protein kinase C, DiC8. It also acted as an inhibitor of [Ca2+]cyt. oscillations. We investigated how these observations could be reconciled with our previously introduced model of [Ca2+]cyt. oscillations in hepatocytes [Somogyi and Stucki (1991) J. Biol. Chem. 266, 11068-11077]. First of all, the effect of calmodulin inhibitors (calmidazolium and CGS 9343 B), acting at the core of our model on the feedback of Ca2+ on Ins(1,4,5)P3-induced Ca2+ release, was not altered by the new modulators. In addition, all agonists and antagonists could be used interchangeably in combination and introduced no significant change in the oscillatory pattern or spike shape. Since the response was solely limited to frequency modulation, over- or understimulation of the oscillatory system, there is no need to create a new oscillator or to introduce further reaction steps into the core of the model. We conclude that the regulation of [Ca2+]cyt. via the explored second-messenger pathways can be embedded into the oscillatory system as modulation of rate constants already present in this model.

SUBMITTER: Somogyi R

PROVIDER: S-EPMC1132985 | biostudies-other | 1992 Sep

REPOSITORIES: biostudies-other

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Project description:The potential for complex synergistic or antagonistic interactions between multiple stressors presents one of the largest uncertainties when predicting ecological change but, despite common use of the terms in the scientific literature, a consensus on their operational definition is still lacking. The identification of synergism or antagonism is generally straightforward when stressors operate in the same direction, but if individual stressor effects oppose each other, the definition of synergism is paradoxical because what is synergistic to one stressor's effect direction is antagonistic to the others. In their highly cited meta-analysis, Crain et al. (Ecology Letters, 11, 2008: 1304) assumed in situations with opposing individual effects that synergy only occurs when the cumulative effect is more negative than the additive sum of the opposing individual effects. We argue against this and propose a new systematic classification based on an additive effects model that combines the magnitude and response direction of the cumulative effect and the interaction effect. A new class of "mitigating synergism" is identified, where cumulative effects are reversed and enhanced. We applied our directional classification to the dataset compiled by Crain et al. (Ecology Letters, 11, 2008: 1304) to determine the prevalence of synergistic, antagonistic, and additive interactions. Compared to their original analysis, we report differences in the representation of interaction classes by interaction type and we document examples of mitigating synergism, highlighting the importance of incorporating individual stressor effect directions in the determination of synergisms and antagonisms. This is particularly pertinent given a general bias in ecology toward investigating and reporting adverse multiple stressor effects (double negative). We emphasize the need for reconsideration by the ecological community of the interpretation of synergism and antagonism in situations where individual stressor effects oppose each other or where cumulative effects are reversed and enhanced.

Project description:BACKGROUND: Understanding the key elements of signaling of chondroprogenitor cells at the earliest steps of differentiation may substantially improve our opportunities for the application of mesenchymal stem cells in cartilage tissue engineering, which is a promising approach of regenerative therapy of joint diseases. Ion channels, membrane potential and Ca(2+)-signaling are important regulators of cell proliferation and differentiation. Our aim was to identify such plasma membrane ion channels involved in signaling during chondrogenesis, which may serve as specific molecular targets for influencing chondrogenic differentiation and ultimately cartilage formation. METHODOLOGY/PRINCIPAL FINDINGS: Using patch-clamp, RT-PCR and Western-blot experiments, we found that chondrogenic cells in primary micromass cell cultures obtained from embryonic chicken limb buds expressed voltage-gated Na(V)1.4, K(V)1.1, K(V)1.3 and K(V)4.1 channels, although K(V)1.3 was not detectable in the plasma membrane. Tetrodotoxin (TTX), the inhibitor of Na(V)1.4 channels, had no effect on cartilage formation. In contrast, presence of 20 mM of the K(+) channel blocker tetraethyl-ammonium (TEA) during the time-window of the final commitment of chondrogenic cells reduced K(V) currents (to 27±3% of control), cell proliferation (thymidine incorporation: to 39±4.4% of control), expression of cartilage-specific genes and consequently, cartilage formation (metachromasia: to 18.0±6.4% of control) and also depolarized the membrane potential (by 9.3±2.1 mV). High-frequency Ca(2+)-oscillations were also suppressed by 10 mM TEA (confocal microscopy: frequency to 8.5±2.6% of the control). Peak expression of TEA-sensitive K(V)1.1 in the plasma membrane overlapped with this period. Application of TEA to differentiated chondrocytes, mainly expressing the TEA-insensitive K(V)4.1 did not affect cartilage formation. CONCLUSIONS/SIGNIFICANCE: These data demonstrate that the differentiation and proliferation of chondrogenic cells depend on rapid Ca(2+)-oscillations, which are modulated by K(V)-driven membrane potential changes. K(V)1.1 function seems especially critical during the final commitment period. We show the critical role of voltage-gated cation channels in the differentiation of non-excitable cells with potential therapeutic use.

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