No abstract

Rett syndrome caused by MeCP2 mutations is a devastating neurodevelopmental disorder accompanied by severe breathing irregularities. Using transduction of organotypic slices from model MeCP2-/y mice with neuron-specific calcium sensor protein D3cpv, we examined the slow calcium buffering in neurons in pre-Bötzinger complex (preBötC), a component of the complex respiratory network. Examination of wild-type (WT) and MeCP2 null mice showed clear differences in the spatial organisations of neurons in preBötC and also in the disturbances in calcium homeostasis in mutant mice during early postnatal development. Deregulated calcium buffering in MeCP2-/y neurons was indicated by increased amplitude and kinetics of depolarisation-induced calcium transients. Both effects were related to an insufficient calcium uptake into the endoplasmic reticulum that was restored after pretreatment with brain-derived neurotrophic factor (BNDF). Conversely, the inhibition of BDNF signalling in WT neurons produced disturbances similar to those observed in MeCP2-/y mice. Brief hypoxia and calcium release from internal stores induced global calcium increases, after which the processes of many MeCP2-/y neurons were retracted, an effect that was also corrected by pretreatment with BDNF. The data obtained point to a tight connection between calcium homeostasis and long-term changes in neuronal connectivity. We therefore propose that calcium-dependent retraction of neurites in preBötC neurons can cause remodelling of the neuronal network during development and set up the conditions for appearance of breathing irregularities in Rett model mice.

Background: cAMP is an ubiquitous second messenger mediating various neuronal functions, often as a consequence of increased intracellular Ca 2+ levels. While imaging of calcium is commonly used in neuroscience applications, probing for cAMP levels has not yet been performed in living vertebrate neuronal tissue before.

J. Neurochem. (2011) 117, 295–308. Abstract The exchange factor directly activated by cAMP (Epac) can couple cAMP production to the activation of particular membrane and cytoplasmic targets. Using patch‐clamp recordings and calcium imaging in organotypic brainstem slices, we examined the role of Epac in pre‐Bötzinger complex, an essential part of the respiratory network. The selective agonist 8‐(4‐chlorophenylthio)‐2′‐O‐methyl‐cAMP (8‐pCPT) sensitized calcium mobilisation from inositol‐1,4,5‐trisphosphate‐sensitive internal stores that stimulated TRPM4 (transient receptor potential cation channel, subfamily M, Melastatin) channels and potentiated the bursts of action potentials. 8‐pCPT actions were abolished after inhibition of phospholipase C with U73122 and depletion of calcium stores with thapsigargin. Caffeine‐sensitive release channels were not modulated by 8‐pCPT. Epac inhibited ATP‐sensitive K+ channels that also led to the enhancement of bursting by 8‐pCPT. Bursting activity, spontaneous calcium transients and activity of TRPM4 and ATP‐sensitive K+ channels were potentiated after brief exposures to bradykinin and incubation with wortmannin produced opposite effects that can be explained by changes in phosphatidylinositol 4,5‐bisphosphate levels. 8‐pCPT stimulated the respiratory motor output in functionally intact preparations and the effects of bradykinin and wortmannin were identical to those observed in organotypic slices. The data thus indicate a novel pathway of controlling bursting activity in pre‐Bötzinger complex neurons through Epac that can involved in reinforcement of the respiratory activity by cAMP.