The Role of Calcium/Calmodulin-Activated Calcineurin in Rapid and Slow Endocytosis at Central Synapses

National Institute of Neurological Disorders and Stroke, Bethesda, Maryland 20892, USA.
The Journal of Neuroscience : The Official Journal of the Society for Neuroscience (Impact Factor: 6.34). 09/2010; 30(35):11838-47. DOI: 10.1523/JNEUROSCI.1481-10.2010
Source: PubMed


Although the calcium/calmodulin-activated phosphatase calcineurin may dephosphorylate many endocytic proteins, it is not considered a key molecule in mediating the major forms of endocytosis at synapses-slow, clathrin-dependent and the rapid, clathrin-independent endocytosis. Here we studied the role of calcineurin in endocytosis by reducing calcium influx, inhibiting calmodulin with pharmacological blockers and knockdown of calmodulin, and by inhibiting calcineurin with pharmacological blockers and knock-out of calcineurin. These manipulations significantly inhibited both rapid and slow endocytosis at the large calyx-type synapse in 7- to 10-d-old rats and mice, and slow, clathrin-dependent endocytosis at the conventional cultured hippocampal synapse of rats and mice. These results suggest that calcium influx during nerve firing activates calcium/calmodulin-dependent calcineurin, which controls the speed of both rapid and slow endocytosis at synapses by dephosphorylating endocytic proteins. The calcium/calmodulin/calcineurin signaling pathway may underlie regulation of endocytosis by nerve activity and calcium as reported at many synapses over the last several decades.

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Available from: Jianhua xu, Mar 30, 2015
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    • "deling study investigates the effects of Ca 2+ buffering by CaM on AP - evoked synaptic vesicle release and short - term synaptic plasticity . The multiple roles of CaM in modulating synaptic transmission , which it exerts via interactions with its target proteins , have been extensively characterized ( Xia and Storm , 2005 ; Pang et al . , 2010 ; Sun et al . , 2010 ; Lipstein et al . ,"
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    ABSTRACT: Action potential-dependent release of synaptic vesicles and short-term synaptic plasticity are dynamically regulated by the endogenous Ca2+ buffers that shape [Ca2+] profiles within a presynaptic bouton. Calmodulin is one of the most abundant presynaptic proteins and it binds Ca2+ faster than any other characterized endogenous neuronal Ca2+ buffer. Direct effects of calmodulin on fast presynaptic Ca2+ dynamics and vesicular release however have not been studied in detail. Using experimentally constrained three-dimensional diffusion modeling of Ca2+ influx–exocytosis coupling at small excitatory synapses we show that, at physiologically relevant concentrations, Ca2+ buffering by calmodulin plays a dominant role in inhibiting vesicular release and in modulating short-term synaptic plasticity. We also propose a novel and potentially powerful mechanism for short-term facilitation based on Ca2+-dependent dynamic dislocation of calmodulin molecules from the plasma membrane within the active zone.
    Frontiers in Cellular Neuroscience 07/2015; 9. DOI:10.3389/fncel.2015.00239 · 4.29 Impact Factor
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    • "Thus, changes in spH fluorescence reflect the relative strength of SV release and recycling (Sankaranarayanan & Ryan, 2000). Because of technical limitation in resolving single action potential (AP)-induced spH responses, we instead used trains of stimulation (20 Hz, 10 s) to provide a sufficient signal-to-noise ratio (Sun et al., 2010). Therefore, applying this dual-channel live imaging enables us to investigate the contribution of motile axonal mitochondria to synaptic function at the single-bouton levels. "
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    ABSTRACT: Mitochondria are cellular power plants that supply ATP to power various biological activities essential for neuronal growth, survival, and function. Due to extremely varied morphological features, neurons face exceptional challenges to maintain energy homeostasis. Neurons require specialized mechanisms distributing mitochondria to distal synapses where energy is in high demand. Axons and synapses undergo activity-dependent remodeling, thereby altering mitochondrial distribution. The uniform microtubule polarity has made axons particularly useful for exploring mechanisms regulating mitochondrial transport. Mitochondria alter their motility under stress conditions or when their integrity is impaired. Therefore, research into the mechanisms regulating mitochondrial motility in healthy and diseased neurons is an important emerging frontier in neurobiology. In this chapter, we discuss the current protocols in the characterization of axonal mitochondrial transport in primary neuron cultures isolated from embryonic rats and adult mice. We also briefly discuss new procedures developed in our lab in analyzing mitochondrial motility patterns at presynaptic terminals and evaluate their impact on synaptic vesicle release.
    Methods in enzymology 11/2014; 547:75-96. DOI:10.1016/B978-0-12-801415-8.00005-9 · 2.09 Impact Factor
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    • "With the active MLCK peptide, the t 50 was 301 ± 53 ms (n = 7; P = 0.89). Endocytosis in cones does not require dynamin and is much faster than in other cells where endocytosis is regulated by CaM (Wu et al., 2009; Sun et al., 2010; Van Hook and Thoreson, 2012). The finding that CaM did not affect cone endocytosis suggests that the slowed recovery from depression is unlikely to be the result of either reduced supply of vesicles in the cone terminal or inhibited release site clearance. "
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    ABSTRACT: At the first synapse in the vertebrate visual pathway, light-evoked changes in photoreceptor membrane potential alter the rate of glutamate release onto second-order retinal neurons. This process depends on the synaptic ribbon, a specialized structure found at various sensory synapses, to provide a supply of primed vesicles for release. Calcium (Ca(2+)) accelerates the replenishment of vesicles at cone ribbon synapses, but the mechanisms underlying this acceleration and its functional implications for vision are unknown. We studied vesicle replenishment using paired whole-cell recordings of cones and postsynaptic neurons in tiger salamander retinas and found that it involves two kinetic mechanisms, the faster of which was diminished by calmodulin (CaM) inhibitors. We developed an analytical model that can be applied to both conventional and ribbon synapses and showed that vesicle resupply is limited by a simple time constant, τ = 1/(Dρδs), where D is the vesicle diffusion coefficient, δ is the vesicle diameter, ρ is the vesicle density, and s is the probability of vesicle attachment. The combination of electrophysiological measurements, modeling, and total internal reflection fluorescence microscopy of single synaptic vesicles suggested that CaM speeds replenishment by enhancing vesicle attachment to the ribbon. Using electroretinogram and whole-cell recordings of light responses, we found that enhanced replenishment improves the ability of cone synapses to signal darkness after brief flashes of light and enhances the amplitude of responses to higher-frequency stimuli. By accelerating the resupply of vesicles to the ribbon, CaM extends the temporal range of synaptic transmission, allowing cones to transmit higher-frequency visual information to downstream neurons. Thus, the ability of the visual system to encode time-varying stimuli is shaped by the dynamics of vesicle replenishment at photoreceptor synaptic ribbons.
    The Journal of General Physiology 10/2014; 144(5). DOI:10.1085/jgp.201411229 · 4.79 Impact Factor
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