Dynamic remodelling of synapses can occur in the absence of the parent cell body

<p>Abstract</p> <p>Background</p> <p>Retraction of nerve terminals is a characteristic feature of development, injury and insult and may herald many neurodegenerative diseases. Although morphological events have been well characterized, we know relatively little about the nature of the underlying cellular machinery. Evidence suggests a strong local component in determining which neuronal branches and synapses are lost, but a greater understanding of this basic neurological process is required. Here we test the hypothesis that nerve terminals are semi-autonomous and able to rapidly respond to local stimuli in the absence of communication with their parent cell body.</p> <p>Results</p> <p>We used an isolated preparation consisting of distal peripheral nerve stumps, associated nerve terminals and post-synaptic muscle fibres, maintained in-vitro for up to 3 hrs. In this system synapses are intact but the presynaptic nerve terminal is disconnected from its cell soma. In control preparations synapses were stable for extended periods and did not undergo Wallerian degneration. In contrast, addition of purines triggers rapid changes at synapses. Using fluorescence and electron microscopy we observe ultrastructural and gross morphological events consistent with nerve terminal retraction. We find no evidence of Wallerian or Wallerian-like degeneration in these preparations. Pharmacological experiments implicate pre-synaptic P2X7 receptor subunits as key mediators of these events.</p> <p>Conclusion</p> <p>The data presented suggest; first that isolated nerve terminals are able to regulate connectivity independent of signals from the cell body, second that synapses exist in a dynamic state, poised to shift from stability to loss by activating intrinsic mechanisms and molecules, and third that local purines acting at purinergic receptors can trigger these events. A role for ATP receptors in this is not surprising since they are frequently activated during cellular injury, when adenosine tri-phosphate is released from damaged cells. Local control demands that the elements necessary to drive retraction are constitutively present. We hypothesize that pre-existing scaffolds of molecular motors and cytoskeletal proteins could provide the dynamism required to drive such structural changes in nerve terminals in the absence of the cell body.</p

Induction of Cell Stress in Neurons from Transgenic Mice Expressing Yellow Fluorescent Protein: Implications for Neurodegeneration Research

Peer reviewedPublisher PD

Rapid loss of motor nerve terminals following hypoxia–reperfusion injury occurs via mechanisms distinct from classic Wallerian degeneration

Motor nerve terminals are known to be vulnerable to a wide range of pathological stimuli. To further characterize this vulnerability, we have developed a novel model system to examine the response of mouse motor nerve terminals in ex vivo nerve/muscle preparations to 2 h hypoxia followed by 2 h reperfusion. This insult induced a rapid loss of neurofilament and synaptic vesicle protein immunoreactivity at pre-synaptic motor nerve terminals but did not appear to affect post-synaptic endplates or muscle fibres. The severity of nerve terminal loss was dependent on the age of the mouse and muscle type: in 8–12-week-old mice the predominantly fast-twitch lumbrical muscles showed an 82.5% loss, whereas the predominantly slow-twitch muscles transversus abdominis and triangularis sterni showed a 57.8% and 27.2% loss, respectively. This was contrasted with a > 97% loss in the predominantly slow-twitch muscles from 5–6-week-old mice. We have also demonstrated that nerve terminal loss occurs by a mechanism distinct from Wallerian degeneration, as the slow Wallerian degeneration (Wlds) gene did not modify the extent of nerve terminal pathology. Together, these data show that our new model of hypoxia–reperfusion injury is robust and repeatable, that it induces rapid, quantitative changes in motor nerve terminals and that it can be used to further examine the mechanisms regulating nerve terminal vulnerability in response to hypoxia–reperfusion injury

Dynamic remodelling of synapses can occur in the absence of the parent cell body-2

<p><b>Copyright information:</b></p><p>Taken from "Dynamic remodelling of synapses can occur in the absence of the parent cell body"</p><p>http://www.biomedcentral.com/1471-2202/8/79</p><p>BMC Neuroscience 2007;8():79-79.</p><p>Published online 26 Sep 2007</p><p>PMCID:PMC2048966.</p><p></p>urofilament 165 and SV2 (green: axons and terminals), visualised with a Cy2-conjugated secondary antibody and co-stained with TRITC-αBTX (red: muscle endplates), 165 min after a 15 min BzATP (100 μM) pulse. Nerve terminals associated with either fragmented (a), or absent (b) muscle endplates were seen, as were fragmented nerve terminals contacting either apparently normal (c), or fragmented endplates (c, d). Scale bar a, d = 15 μm, b, c = 30 μ

Dynamic remodelling of synapses can occur in the absence of the parent cell body-1

<p><b>Copyright information:</b></p><p>Taken from "Dynamic remodelling of synapses can occur in the absence of the parent cell body"</p><p>http://www.biomedcentral.com/1471-2202/8/79</p><p>BMC Neuroscience 2007;8():79-79.</p><p>Published online 26 Sep 2007</p><p>PMCID:PMC2048966.</p><p></p>plates present in: 300 min control (open bar); 165 min subsequent to a 15 min BzATP pulse (100 μM: filled bar); 165 min subsequent to a 15 min BzATP pulse (filled bar) in the presence of Brilliant Blue G (a selective P2X7 receptor antagonist: BBG, 1 μM: broad diagonal cross-hatching); 165 min subsequent to a 15 min BzATP pulse in the presence of Reactive Blue 2 (which blocks P2Y and P2X receptor subunits: 100 μM, RB2: narrow diagonal cross-hatching) or 165 min subsequent to a 15 min BzATP pulse in the presence of Suramin (a broad spectrum P2 receptor agonist: 100 μM: stipple). All data is mean ± SEM of multiple repeats (see text for n values)

Dynamic remodelling of synapses can occur in the absence of the parent cell body-6

<p><b>Copyright information:</b></p><p>Taken from "Dynamic remodelling of synapses can occur in the absence of the parent cell body"</p><p>http://www.biomedcentral.com/1471-2202/8/79</p><p>BMC Neuroscience 2007;8():79-79.</p><p>Published online 26 Sep 2007</p><p>PMCID:PMC2048966.</p><p></p>urofilament 165 and SV2 (green: axons and terminals), visualised with a Cy2-conjugated secondary antibody and co-stained with TRITC-αBTX (red: muscle endplates), 165 min after a 15 min BzATP pulse (100 μM). Both 'occupied' (a: arrow) and 'unoccupied' (a: arrowhead) endplates were present, as were unoccupied endplates with intact presynaptic axons (b). Many terminals appeared to be in the process of retraction resulting in minor (c: asterisk) or major (d: asterisks) regions of unoccupied endplates. These were classified as 'intermediate' in subsequent data. Control nerve/muscle preparations, which were maintained in physiological saline for up to 300 min, showed no visible signs of degeneration or retraction of nerve terminals (e). Scale bar a-e = 15 μ

Dynamic remodelling of synapses can occur in the absence of the parent cell body-8

<p><b>Copyright information:</b></p><p>Taken from "Dynamic remodelling of synapses can occur in the absence of the parent cell body"</p><p>http://www.biomedcentral.com/1471-2202/8/79</p><p>BMC Neuroscience 2007;8():79-79.</p><p>Published online 26 Sep 2007</p><p>PMCID:PMC2048966.</p><p></p>urofilament 165 and SV2 (green: axons and terminals), visualised with a Cy2-conjugated secondary antibody and co-stained with TRITC-αBTX (red: muscle endplates), 165 min after a 15 min BzATP (100 μM) pulse. Nerve terminals associated with either fragmented (a), or absent (b) muscle endplates were seen, as were fragmented nerve terminals contacting either apparently normal (c), or fragmented endplates (c, d). Scale bar a, d = 15 μm, b, c = 30 μ

Dynamic remodelling of synapses can occur in the absence of the parent cell body-7

<p><b>Copyright information:</b></p><p>Taken from "Dynamic remodelling of synapses can occur in the absence of the parent cell body"</p><p>http://www.biomedcentral.com/1471-2202/8/79</p><p>BMC Neuroscience 2007;8():79-79.</p><p>Published online 26 Sep 2007</p><p>PMCID:PMC2048966.</p><p></p>plates present in: 300 min control (open bar); 165 min subsequent to a 15 min BzATP pulse (100 μM: filled bar); 165 min subsequent to a 15 min BzATP pulse (filled bar) in the presence of Brilliant Blue G (a selective P2X7 receptor antagonist: BBG, 1 μM: broad diagonal cross-hatching); 165 min subsequent to a 15 min BzATP pulse in the presence of Reactive Blue 2 (which blocks P2Y and P2X receptor subunits: 100 μM, RB2: narrow diagonal cross-hatching) or 165 min subsequent to a 15 min BzATP pulse in the presence of Suramin (a broad spectrum P2 receptor agonist: 100 μM: stipple). All data is mean ± SEM of multiple repeats (see text for n values)