Orexin (OX)/hypocretin-containing neurons are main regulators of wakefulness stability, arousal, and energy homeostasis. Their activity varies in relation to the animal's behavioral state. We here tested whether such variation is subserved by synaptic plasticity phenomena in basal conditions. Mice were sacrificed during day or night, at times when sleep or wake, respectively, predominates, as assessed by electroencephalography in matched mice. Triple immunofluorescence was used to visualize OX-A perikarya and varicosities containing the vesicular glutamate transporter (VGluT)2 or the vesicular GABA transporter (VGAT) combined with synaptophysin (Syn) as a presynaptic marker. Appositions on OX-A(+) somata were quantitatively analyzed in pairs of sections in epifluorescence and confocal microscopy. The combined total number of glutamatergic (Syn(+)/VGluT2(+)) and GABAergic (Syn(+)/VGAT(+)) varicosities apposed to OX-A somata was similar during day and night. However, glutamatergic varicosities were significantly more numerous at night, whereas GABAergic varicosities prevailed in the day. Triple immunofluorescence in confocal microscopy was employed to visualize synapse scaffold proteins as postsynaptic markers and confirmed the nighttime prevalence of VGluT2(+) together with postsynaptic density protein 95(+) excitatory contacts, and daytime prevalence of VGAT(+) together with gephyrin(+) inhibitory contacts, while also showing that they formed synapses on OX-A(+) cell bodies. The findings reveal a daily reorganization of axosomatic synapses in orexinergic neurons, with a switch from a prevalence of excitatory innervation at a time corresponding to wakefulness to a prevalence of inhibitory innervations in the antiphase, at a time corresponding to sleep. This reorganization could represent a key mechanism of plasticity of the orexinergic network in basal conditions.
Laperchia, Claudia;Imperatore, Roberta;Azeez, Idris Ayodeji;Del Gallo, Federico;Bertini, Giuseppe;CRISTINO, Luigia;Bentivoglio, Marina
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