In vivo Two-Photon Imaging of Anesthesia-Specific Alterations in Microglial Surveillance and Photodamage-Directed Motility in Mouse Cortex

Two-photon imaging of fluorescently labeled microglia in vivo provides a direct approach to measure motility of microglial processes as a readout of microglial function that is crucial in the context of neurodegenerative diseases, as well as to understand the neuroinflammatory response to implanted substrates and brain-computer interfaces. In this longitudinal study, we quantified surveilling and photodamage-directed microglial processes motility in both acute and chronic cranial window preparations and compared the motility under isoflurane and ketamine anesthesia to an awake condition in the same animal. The isoflurane anesthesia increased the length of surveilling microglial processes in both acute and chronic preparations, while ketamine increased the number of microglial branches in acute preparation only. In chronic (but not acute) preparation, the extension of microglial processes toward the laser-ablated microglial cell was faster under isoflurane (but not ketamine) anesthesia than in awake mice, indicating distinct effects of anesthetics and of preparation type. These data reveal potentiating effects of isoflurane on microglial response to damage, and provide a framework for comparison and optimal selection of experimental conditions for quantitative analysis of microglial function using two-photon microscopy in vivo.Peer reviewe

Opposite Reactivity of Meningeal versus Cortical Microvessels to the Nitric Oxide Donor Glyceryl Trinitrate Evaluated In Vivo with Two-Photon Imaging

Peer reviewe

Longitudinal two-photon imaging in somatosensory cortex of behaving mice reveals dendritic spine formation enhancement by subchronic administration of low-dose ketamine

Ketamine, a well-known anesthetic, has recently attracted renewed attention as a fast-acting antidepressant. A single dose of ketamine induces rapid synaptogenesis, which may underlie its antidepressant effect. To test whether repeated exposure to ketamine triggers sustained synaptogenesis, we administered a sub-anesthetic dose of ketamine (10 mg/kg i.p.) once-daily for 5 days, and repeatedly imaged dendritic spines of the YFP-expressing pyramidal neurons in somatosensory cortex of awake female mice using in vivo two-photon microscopy. We found that the spine formation rate became significantly higher at 72-132 h after the first ketamine injection (but not at 6-24 h), while the rate of elimination of pre-existing spines remained unchanged. In contrast to the net gain of spines observed in ketamine-treated mice, the vehicle-injected control mice exhibited a net loss typical for young-adult animals undergoing synapse pruning. Ketamine-induced spinogenesis was correlated with increased PSD-95 and phosphorylated actin, consistent with formation of new synapses. Moreover, structural synaptic plasticity caused by ketamine was paralleled by a significant improvement in the nest building behavioral assay. Taken together, our data show that subchronic low-dose ketamine induces a sustained shift towards spine formation.Peer reviewe

In vivo Two-Photon Imaging of Anesthesia-Specific Alterations in Microglial Surveillance and Photodamage-Directed Motility in Mouse Cortex

Two-photon imaging of fluorescently labeled microglia in vivo provides a direct approach to measure motility of microglial processes as a readout of microglial function that is crucial in the context of neurodegenerative diseases, as well as to understand the neuroinflammatory response to implanted substrates and brain-computer interfaces. In this longitudinal study, we quantified surveilling and photodamage-directed microglial processes motility in both acute and chronic cranial window preparations and compared the motility under isoflurane and ketamine anesthesia to an awake condition in the same animal. The isoflurane anesthesia increased the length of surveilling microglial processes in both acute and chronic preparations, while ketamine increased the number of microglial branches in acute preparation only. In chronic (but not acute) preparation, the extension of microglial processes toward the laser-ablated microglial cell was faster under isoflurane (but not ketamine) anesthesia than in awake mice, indicating distinct effects of anesthetics and of preparation type. These data reveal potentiating effects of isoflurane on microglial response to damage, and provide a framework for comparison and optimal selection of experimental conditions for quantitative analysis of microglial function using two-photon microscopy in vivo

Superhydrophobic Lubrication

| openaire: EC/H2020/725513/EU//SuperRepelLubrication is one of the most important ways to reduce the effect of friction, which is the single largest cause for energy losses in society. Typically, friction reduction is done by lubrication with petroleum-based oils, while technology focus is shifting toward environmentally-friendly green lubrication. Lowest friction coefficients with water-based lubrication have previously been achieved with smooth surfaces such as silicon carbide and silicon nitride or polyzwitterionic polymer brushes with typical coefficients of friction in the order of 0.002. Here, a novel concept for green lubrication using a bilayer of water and ambient air acting as the lubricant between a hydrophilic and superhydrophobic surface is shown. This method achieves superlubricity with friction coefficients down to 0.002 as measured with oscillating tribometer and tilting stage. In addition, possible applications for superhydrophobic lubrication such as tunable lubrication and a 2D mouse treadmill, are shown.Peer reviewe

Dissociation Between Neuronal and Astrocytic Calcium Activity in Response to Locomotion in Mice

Locomotion triggers a coordinated response of both neurons and astrocytes in the brain. Here we performed calcium (Ca2+) imaging of these two cell types in the somatosensory cortex in head-fixed mice moving on the airlifted platform. Ca2+ activity in astrocytes significantly increased during locomotion from a low quiescence level. Ca2+ signals first appeared in the distal processes and then propagated to astrocytic somata, where it became significantly larger and exhibited oscillatory behaviour. Thus, astrocytic soma operates as both integrator and amplifier of Ca2+ signal. In neurons, Ca2+ activity was pronounced in quiescent periods and further increased during locomotion. Neuronal Ca2+ concentration ([Ca2+]i) rose almost immediately following the onset of locomotion, whereas astrocytic Ca2+ signals lagged by several seconds. Such a long lag suggests that astrocytic [Ca2+]i elevations are unlikely to be triggered by the activity of synapses among local neurons. Ca2+ responses to pairs of consecutive episodes of locomotion did not significantly differ in neurons, while were significantly diminished in response to the second locomotion in astrocytes. Such astrocytic refractoriness may arise from distinct mechanisms underlying Ca2+ signal generation. In neurons, the bulk of Ca2+ enters through the Ca2+ channels in the plasma membrane allowing for steady-level Ca2+ elevations in repetitive runs. Astrocytic Ca2+ responses originate from the intracellular stores, the depletion of which affects subsequent Ca2+ signals. Functionally, neuronal Ca2+ response reflects sensory input processed by neurons. Astrocytic Ca2+ dynamics is likely to provide metabolic and homeostatic support within the brain active milieu.</p

The action of of GTN on meningeal vessels in open cranial window preparation.

<p><b>A</b>, examples of meningeal vessels before (upper image) and after GTN administration (lower image). <b>B</b>, the normalized diameter of meningeal arterioles after injection of the GTN (filled circles) or vehicle (empty circles), real diameter before application 17.4±2 µm for meningeal arterioles and 22.8±4 µm for meningeal venules. <b>C</b>, comparison of changes in the area under curve in GTN vs vehicle in meningeal arterioles (n = 6 and n = 4, respectively, ***  =  P<0.001), <b>D</b> and <b>E</b>, the same for meningeal venules (n = 5 and n = 4, respectively). Note that GTN changed the diameter of arterioles but not venules. Scale bar 40 µm.</p

The action of L-NAME (filled circles) and vehicle (open circles) on the diameter of cortical and meningeal vessels in open cranial window preparation.

<p>A, the time of changes in cortical arterioles (real diameter 38.1±6 µm), B, cortical venules (real diameter 46.4±13 µm), D, meningeal arterioles (real diameter 20.7±5 µm) and E, meningeal venules (real diameter 24.1±7 µm), respectively. C and F, quantification of results (n = 9, 4, 8, 8 for meningeal arterioles, meningeal venules, cortical arterioles and cortical venules, respectively, *** = P<0.001, ** = P<0.01). Notice constriction of cortical but not of meningeal vessels.</p

Schematic presentation of potential mechanisms for opposite modulation of dura mater and pial/cortical vessels by GTN.

<p>In <i>dura mater</i> occupied by mast cells and densely innervated by trigeminal and autonomous nerves GTN can induce vasoconstriction of small vessels either directly, or via release of vasoconstrictory agents from mast cells or through the neuronal control. The functional outcome depends on the combination of vasodilatatory versus vasoconstrictory agents and on the receptor profile. The dilatatory effect of GTN in pial/cortical vessels could be due to the direct action of this agent on the vessel wall or mediated via astrocytes releasing NO.</p

Effects of GTN on cortical vessels in the open cranial window preparation.

<p>A, examples of cortical vessels before (upper image) and after GTN administration (lower image). B, the normalized diameter of cortical arterioles after injection of the GTN (filled circles) or vehicle (empty circles), real diameter before application 36.1±6 µm for cortical arterioles and 38±8 µm for cortical venules. C, comparison of changes in the area under curve (AUC) in GTN versus vehicle in cortical arterioles (n = 6 and n = 3, respectively). D and E, the same for cortical venules (n = 8 and n = 4, respectively). Note that GTN significantly changed the diameter of arterioles (*  =  P<0.05) but not venules (P = 0.09). Scale bar 25 µm.</p